A phylogenomic study of endosymbiotic bacteria

Endosymbiotic bacteria of aphids, Buchnera aphidicola, and tsetse flies, Wigglesworthia glossinidia, are descendents of free-living gamma-Proteobacteria. The acceleration of sequence evolution in the endosymbiont genomes is here estimated from a phylogenomic analysis of the gamma-Proteobacteria. The tree topologies associated with the most highly conserved genes suggest that the endosymbionts form a sister group with Escherichia coli, Salmonella sp., and Yersinia pestis. Our results indicate that deviant tree topologies result from high substitution rates and biased nucleotide patterns, rather than from lateral gene transfer, as previously suggested. A reinvestigation of the relative rate increase in the endosymbiont genomes reveals variability among genes that correlate with host-associated metabolic dependencies. The conclusion is that host-level selection has retarded both the loss of genes and the acceleration of sequence evolution in endocellular symbionts.     

Assessment of phylogenomic and orthology approaches for phylogenetic inference

MOTIVATION: Phylogenomics integrates the vast amount of phylogenetic information contained in complete genome sequences, and is rapidly becoming the standard for reliably inferring species phylogenies. There are, however, fundamental differences between the ways in which phylogenomic approaches like gene content, superalignment, superdistance and supertree integrate the phylogenetic information from separate orthologous groups. Furthermore, they all depend on the method by which the orthologous groups are initially determined. Here, we systematically compare these four phylogenomic approaches, in parallel with three approaches for large-scale orthology determination: pairwise orthology, cluster orthology and tree-based orthology. RESULTS: Including various phylogenetic methods, we apply a total of 54 fully automated phylogenomic procedures to the fungi, the eukaryotic clade with the largest number of sequenced genomes, for which we retrieved a golden standard phylogeny from the literature. Phylogenomic trees based on gene content show, relative to the other methods, a bias in the tree topology that parallels convergence in lifestyle among the species compared, indicating convergence in gene content. CONCLUSIONS: Complete genomes are no guarantee for good or even consistent phylogenies. However, the large amounts of data in genomes enable us to carefully select the data most suitable for phylogenomic inference. In terms of performance, the superalignment approach, combined with restrictive orthology, is the most successful in recovering a fungal phylogeny that agrees with current taxonomic views, and allows us to obtain a high-resolution phylogeny. We provide solid support for what has grown to be a common practice in phylogenomics during its advance in recent years. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.     

The evolutionary origin of Xanthomonadales genomes and the nature of the horizontal gene transfer process

Determining the influence of horizontal gene transfer (HGT) on phylogenomic analyses and the retrieval of a tree of life is relevant for our understanding of microbial genome evolution. It is particularly difficult to differentiate between phylogenetic incongruence due to noise and that resulting from HGT. We have performed a large-scale, detailed evolutionary analysis of the different phylogenetic signals present in the genomes of Xanthomonadales, a group of Proteobacteria. We show that the presence of phylogenetic noise is not an obstacle to infer past and present HGTs during their evolution. The scenario derived from this analysis and other recently published reports reflect the confounding effects on bacterial phylogenomics of past and present HGT. Although transfers between closely related species are difficult to detect in genome-scale phylogenetic analyses, past transfers to the ancestor of extant groups appear as conflicting signals that occasionally might make impossible to determine the evolutionary origin of the whole genome.     

Addressing inter-gene heterogeneity in maximum likelihood phylogenomic analysis: yeasts revisited

Phylogenomic approaches to the resolution of inter-species relationships have become well established in recent years. Often these involve concatenation of many orthologous genes found in the respective genomes followed by analysis using standard phylogenetic models. Genome-scale data promise increased resolution by minimising sampling error, yet are associated with well-known but often inappropriately addressed caveats arising through data heterogeneity and model violation. These can lead to the reconstruction of highly-supported but incorrect topologies. With the aim of obtaining a species tree for 18 species within the ascomycetous yeasts, we have investigated the use of appropriate evolutionary models to address inter-gene heterogeneities and the scalability and validity of supermatrix analysis as the phylogenetic problem becomes more difficult and the number of genes analysed approaches truly phylogenomic dimensions. We have extended a widely-known early phylogenomic study of yeasts by adding additional species to increase diversity and augmenting the number of genes under analysis. We have investigated sophisticated maximum likelihood analyses, considering not only a concatenated version of the data but also partitioned models where each gene constitutes a partition and parameters are free to vary between the different partitions (thereby accounting for variation in the evolutionary processes at different loci). We find considerable increases in likelihood using these complex models, arguing for the need for appropriate models when analyzing phylogenomic data. Using these methods, we were able to reconstruct a well-supported tree for 18 ascomycetous yeasts spanning about 250 million years of evolution.     

Bacterial endosymbionts of insects: insights from comparative genomics

The development of molecular techniques for the study of uncultured bacteria allowed the extensive study of the widespread association between insects and intracellular symbiotic bacteria. Most of the bacterial endosymbionts involved in such associations are gamma-proteobacteria, closely related to Escherichia coli. In recent years, five genomes from insect endosymbionts have been sequenced, allowing the performance of extensive genome comparative analysis that, as a complement of phylogenetic studies, and analysis on individual genes, can help to understand the different traits of this particular association, including how the symbiotic process is established, the explanation of the special features of these microbial genomes, the bases of this intimate association and the possible future that awaits the endosymbionts with extremely reduced genomes.     

Identification of the mammalian Not gene via a phylogenomic approach

Despite the great morphological diversity of early embryos, the underlying mechanisms of gastrulation are known to be broadly conserved in vertebrates. However, a number of genes characterized as fulfilling an essential function in this process in several model organisms display no clear ortholog in mammalian genomes. We have devised an in silico phylogenomic approach, based on exhaustive similarity searches in vertebrate genomes and subsequent bayesian phylogenetic analyses, to identify such missing genes, presumed to be highly divergent. This approach has been used to identify mammalian orthologs of Not, an homeodomain containing gene previously characterized in Xenopus, chick and zebrafish as playing a critical role in the formation of the notochord. This attempt led to the identification of a highly divergent mammalian Not-related gene in the mouse, human and rat. The results from phylogenetic reconstructions, synteny analyses, expression pattern analyses in wild-type and mutant mouse embryos, and overexpression experiments in Xenopus embryos converge to confirm these genes as representatives of the Not family in mammals. The identification of the mammalian Not gene delivers an important component for the understanding of the genetics underlying notochord formation in mammals and its evolution among vertebrates. The phylogenomic method used to retrieve this gene thus provides a tool, which can complement or validate genome annotations in situations when they are weakly supported.     

A congruent solution to arthropod phylogeny: phylogenomics,microRNAs and morphology support monophyletic Mandibulata

While a unique origin of the euarthropods is well established, relationships between the four euarthropod classes—chelicerates, myriapods, crustaceans and hexapods—are less clear. Unsolved questions include the position of myriapods, the monophyletic origin of chelicerates, and the validity of the close relationship of euarthropods to tardigrades and onychophorans. Morphology predicts that myriapods, insects and crustaceans form a monophyletic group, the Mandibulata, which has been contradicted by many molecular studies that support an alternative Myriochelata hypothesis (Myriapoda plus Chelicerata). Because of the conflicting insights from published molecular datasets, evidence from nuclear-coding genes needs corroboration from independent data to define the relationships among major nodes in the euarthropod tree. Here, we address this issue by analysing two independent molecular datasets: a phylogenomic dataset of 198 protein-coding genes including new sequences for myriapods, and novel microRNA complements sampled from all major arthropod lineages. Our phylogenomic analyses strongly support Mandibulata, and show that Myriochelata is a tree-reconstruction artefact caused by saturation and long-branch attraction. The analysis of the microRNA dataset corroborates the Mandibulata, showing that the microRNAs miR-965 and miR-282 are present and expressed in all mandibulate species sampled, but not in the chelicerates. Mandibulata is further supported by the phylogenetic analysis of a comprehensive morphological dataset covering living and fossil arthropods, and including recently proposed, putative apomorphies of Myriochelata. Our phylogenomic analyses also provide strong support for the inclusion of pycnogonids in a monophyletic Chelicerata, a paraphyletic Cycloneuralia, and a common origin of Arthropoda (tardigrades, onychophorans and arthropods), suggesting that previous phylogenies grouping tardigrades and nematodes may also have been subject to tree-reconstruction artefacts.     

How well do multispecies coalescent methods perform with mitochondrial genomic data? A case study of butterflies and moths (Insecta: Lepidoptera)

Despite the broad adoption of multispecies coalescent (MSC) methods for nuclear phylogenomics, they have yet to be applied to mitochondrial (mt) genomic data. As the potential sources of phylogenomic bias that MSC methods can address, such as incomplete lineage sorting, horizontal gene transfer and gene tree heterogeneity, have been found in mt genomic data, these approaches may improve the accuracy of phylogenetic inference with these data. In the present study, we examined the behaviour of MSC methods in reconstructing the phylogeny of Lepidoptera (butterflies and moths), a group for which mt genomic data are known to have strong resolving power. Traditional concatenation methods of analysing mt genomes for Lepidoptera infer topologies highly congruent with those generated from independent nuclear datasets. Individual mt gene trees performed poorly in recovering consensus relationships at deep levels (i.e. superfamily monophyly and inter-relationships) and only moderately well for shallow relationships (i.e. within Papilionoidea). In contrast, MSC analyses with ASTRAL performed strongly with almost complete concordance to both concatenated mt genome analyses and independent nuclear analyses at both deep and shallow phylogenetic scales. Outgroup choice had a limited impact on tree accuracy, with even phylogenetically distant outgroups still resulting in topologies highly congruent with results from nuclear datasets, although MSC analyses appeared to be marginally more affected by outgroup choice than concatenation analyses. In general, discordance between concatenation and MSC analyses was found at nodes whose resolution varied between previous nuclear phylogenomic studies. The sensitivity of individual relationships to analysis with MSC vs concatenation can thus be used to test the robustness of phylogenetic hypotheses. For insect phylogenetics, MSC is a reliable inference method for mt genomic data and is thus a useful complement to the already widely used concatenation approaches.     

Comparative genomics of microbial pathogens and symbionts

We are interested in quantifying the contribution of gene acquisition, loss, expansion and rearrangements to the evolution of microbial genomes. Here, we discuss factors influencing microbial genome divergence based on pair-wise genome comparisons of closely related strains and species with different lifestyles. A particular focus is on intracellular pathogens and symbionts of the genera Rickettsia, Bartonella and BUCHNERA: Extensive gene loss and restricted access to phage and plasmid pools may provide an explanation for why single host pathogens are normally less successful than multihost pathogens. We note that species-specific genes tend to be shorter than orthologous genes, suggesting that a fraction of these may represent fossil-orfs, as also supported by multiple sequence alignments among species. The results of our genome comparisons are placed in the context of phylogenomic analyses of alpha and gamma proteobacteria. We highlight artefacts caused by different rates and patterns of mutations, suggesting that atypical phylogenetic placements can not a priori be taken as evidence for horizontal gene transfer events. The flexibility in genome structure among free-living microbes contrasts with the extreme stability observed for the small genomes of aphid endosymbionts, in which no rearrangements or inflow of genetic material have occurred during the past 50 millions years (1). Taken together, the results suggest that genomic stability correlate with the content of repeated sequences and mobile genetic elements, and thereby indirectly with bacterial lifestyles.     

The First Mitochondrial Genome for Caddisfly (Insecta: Trichoptera) with Phylogenetic Implications

The Trichoptera (caddisflies) is a holometabolous insect order with 14,300 described species forming the second most species-rich monophyletic group of animals in freshwater. Hitherto, there is no mitochondrial genome reported of this order. Herein, we describe the complete mitochondrial (mt) genome of a caddisfly species, Eubasilissa regina (McLachlan, 1871). A phylogenomic analysis was carried out based on the mt genomic sequences of 13 mt protein coding genes (PCGs) and two rRNA genes of 24 species belonging to eight holometabolous orders. Both maximum likelihood and Bayesian inference analyses highly support the sister relationship between Trichoptera and Lepidoptera.     

土蜂科线粒体基因组序列测定和分析

【目的】本研究旨在通过针尾部(Aculeata)昆虫线粒体基因组系统发育分析认知土蜂科(Scoliidae)的单系性及系统发育位置。【方法】利用Illumina Hiseq2500二代测序技术测序土蜂科3属5种的线粒体基因组,并进行注释和分析;基于针尾部昆虫36个线粒体基因组13个蛋白质编码基因(protein-coding genes, PCGs)和2个rRNA基因序列采用最大似然法(maximum likelihood, ML)和贝叶斯法(Bayesian inference, BI)法构建系统发育树。【结果】新测序的土蜂科5个线粒体基因组为五带波壁土蜂Colpa quinquecincta线粒体基因组(GenBank登录号：OM103696),齿石波壁土蜂Colpa tartara线粒体基因组(GenBank登录号：OM103697),厚大长腹土蜂Megacampsomeris grossa线粒体基因组(GenBank登录号：OM103796),台湾大长腹土蜂Megacampsomeris formosensis线粒体基因组(GenBank登录号：OM142776)和斯式土蜂Sc...     

A consistent phylogenetic backbone for the fungi

The kingdom of fungi provides model organisms for biotechnology, cell biology, genetics, and life sciences in general. Only when their phylogenetic relationships are stably resolved, can individual results from fungal research be integrated into a holistic picture of biology. However, and despite recent progress, many deep relationships within the fungi remain unclear. Here, we present the first phylogenomic study of an entire eukaryotic kingdom that uses a consistency criterion to strengthen phylogenetic conclusions. We reason that branches (splits) recovered with independent data and different tree reconstruction methods are likely to reflect true evolutionary relationships. Two complementary phylogenomic data sets based on 99 fungal genomes and 109 fungal expressed sequence tag (EST) sets analyzed with four different tree reconstruction methods shed light from different angles on the fungal tree of life. Eleven additional data sets address specifically the phylogenetic position of Blastocladiomycota, Ustilaginomycotina, and Dothideomycetes, respectively. The combined evidence from the resulting trees supports the deep-level stability of the fungal groups toward a comprehensive natural system of the fungi. In addition, our analysis reveals methodologically interesting aspects. Enrichment for EST encoded data-a common practice in phylogenomic analyses-introduces a strong bias toward slowly evolving and functionally correlated genes. Consequently, the generalization of phylogenomic data sets as collections of randomly selected genes cannot be taken for granted. A thorough characterization of the data to assess possible influences on the tree reconstruction should therefore become a standard in phylogenomic analyses.     

Phylogenetic analysis of C, M, N, and U genomes and their relationships with Triticum and other related genomes as revealed by LMW-GS genes at Glu-3 loci

Phylogenetic relationships between the C, U, N, and M genomes of Aegilops species and the genomes of common wheat and other related species were investigated by using three types of low-molecular-weight glutenin subunit (LMW-GS) genes at Glu-3 loci. A total of 20 LMW-GS genes from Aegilops and Triticum species were isolated, including 11 LMW-m type and 9 LMW-i type genes. Particularly, four LMW-m type and three LMW-i type subunits encoded by the genes on the C, N, and U genomes possessed an extra cysteine residue at conserved positions, which could provide useful information for understanding phylogenetic relationships among Aegilops and Triticum genomes. Phylogenetic trees constructed by using either LMW-i or the combination of LMW-m and LMW-s, as well as analysis of all the three types of LMW-GS genes together, demonstrated that the C and U genomes were closely related to the A genome, whereas the N and M genomes were closely related to the D genome. Our results support previous findings that the A genome was derived from Triticum uratu, the B genome was from Aegilops speltoides, and the D genome was from Aegilops tauschii. In addition, phylogenetic relationships among different genomes analysed in this study support the concept that Aegilops is not monophyletic.     

Evolution of Buchlo? dactyloides based on cloning and sequencing of matK, rbcL, and cob genes from plastid and mitochondrial genomes.

Buffalograss (Buchlo? dactyloides (Nutt.) Englem), a C4 turfgrass species, is native to the Great Plains region of North America. The evolutionary implications of buffalograss are unclear. Sequencing of rbcL and matK genes from plastid and the cob gene from mitochondrial genomes was examined to elucidate buffalo grass evolution. This study is the first to report sequencing of these genes from organelle genomes in the genus Buchlo?. Comparisons of sequence data from the mitochondrial and plastid genome revealed that all genotypes contained the same cytoplasmic origin. There were some rearrangements detected in mitochondrial genome. The buffalograss genome appears to have evolved through the rearrangements of convergent subgenomic domains. Combined analyses of plastid genes suggest that the evolutionary process in Buchlo? accessions studied was monophyletic rather than polyphyletic. However, since plastid and mitochondrial genomes are generally uniparentally inherited, the evolutionary history of these genomes may not reflect the evolutionary history of the organism, especially in a species in which out-crossing is common. The sequence information obtained from this study can be used as a genome-specific marker for investigation of the buffalograss polyploidy complex and testing of the mode of plastid and mitochondrial transmission in genus Buchlo?.     

Computational methods for Gene Orthology inference

Accurate inference of orthologous genes is a pre-requisite for most comparative genomics studies, and is also important for functional annotation of new genomes. Identification of orthologous gene sets typically involves phylogenetic tree analysis, heuristic algorithms based on sequence conservation, synteny analysis, or some combination of these approaches. The most direct tree-based methods typically rely on the comparison of an individual gene tree with a species tree. Once the two trees are accurately constructed, orthologs are straightforwardly identified by the definition of orthology as those homologs that are related by speciation, rather than gene duplication, at their most recent point of origin. Although ideal for the purpose of orthology identification in principle, phylogenetic trees are computationally expensive to construct for large numbers of genes and genomes, and they often contain errors, especially at large evolutionary distances. Moreover, in many organisms, in particular prokaryotes and viruses, evolution does not appear to have followed a simple 'tree-like' mode, which makes conventional tree reconciliation inapplicable. Other, heuristic methods identify probable orthologs as the closest homologous pairs or groups of genes in a set of organisms. These approaches are faster and easier to automate than tree-based methods, with efficient implementations provided by graph-theoretical algorithms enabling comparisons of thousands of genomes. Comparisons of these two approaches show that, despite conceptual differences, they produce similar sets of orthologs, especially at short evolutionary distances. Synteny also can aid in identification of orthologs. Often, tree-based, sequence similarity- and synteny-based approaches can be combined into flexible hybrid methods.     

A mitochondrial genome phylogeny of the Neuropterida (lace-wings, alderflies and snakeflies) and their relationship to the other holometabolous insect orders

We present a mitochondrial (mt) genome phylogeny inferring relationships within Neuropterida (lacewings, alderflies and camel flies) and between Neuropterida and other holometabolous insect orders. Whole mt genomes were sequenced for Sialis hamata (Megaloptera: Sialidae), Ditaxis latistyla (Neuroptera: Mantispidae), Mongoloraphidia harmandi (Raphidioptera: Raphidiidae), Macrogyrus oblongus (Coleoptera: Gyrinidae), Rhopaea magnicornis (Coleoptera: Scarabaeidae), and Mordella atrata (Coleoptera: Mordellidae) and compared against representatives of other holometabolous orders in phylogenetic analyses. Additionally, we test the sensitivity of phylogenetic inferences to four analytical approaches: inclusion vs. exclusion of RNA genes, manual vs. algorithmic alignments, arbitrary vs. algorithmic approaches to excluding variable gene regions and how each approach interacts with phylogenetic inference methods (parsimony vs. Bayesian inference). Of these factors, phylogenetic inference method had the most influence on interordinal relationships. Bayesian analyses inferred topologies largely congruent with morphologically‐based hypotheses of neuropterid relationships, a monophyletic Neuropterida whose sister group is Coleoptera. In contrast, parsimony analyses failed to support a monophyletic Neuropterida as Raphidioptera was the sister group of the entire Holometabola excluding Hymenoptera, and Neuroptera + Megaloptera is the sister group of Diptera, a relationship which has not previously been proposed based on either molecular or morphological data sets. These differences between analytical methods are due to the high among site rate heterogeneity found in insect mt genomes which is properly modelled by Bayesian methods but results in artifactual relationships under parsimony. Properly analysed, the mt genomic data set presented here is among the first molecular data to support traditional, morphology‐based interpretations of relationships between the three neuropterid orders and their grouping with Coleoptera.     

Tree of life based on genome context networks

Efforts in phylogenomics have greatly improved our understanding of the backbone tree of life. However, due to the systematic error in sequence data, a sequence-based phylogenomic approach leads to well-resolved but statistically significant incongruence. Thus, independent test of current phylogenetic knowledge is required. Here, we have devised a distance-based strategy to reconstruct a highly resolved backbone tree of life, on the basis of the genome context networks of 195 fully sequenced representative species. Along with strongly supporting the monophylies of three superkingdoms and most taxonomic sub-divisions, the derived tree also suggests some intriguing results, such as high G+C gram positive origin of Bacteria, classification of Symbiobacterium thermophilum and Alcanivorax borkumensis in Firmicutes. Furthermore, simulation analyses indicate that addition of more gene relationships with high accuracy can greatly improve the resolution of the phylogenetic tree. Our results demonstrate the feasibility of the reconstruction of highly resolved phylogenetic tree with extensible gene networks across all three domains of life. This strategy also implies that the relationships between the genes (gene network) can define what kind of species it is.