Tree Genome Research Articles

A flexible multi-species genome-wide 60K SNP chip developed from pooled resequencing of 240 Eucalyptus tree genomes across 12 species.

We used whole genome resequencing of pooled individuals to develop a high-density single-nucleotide polymorphism (SNP) chip for Eucalyptus. Genomes of 240 trees of 12 species were sequenced at 3.5× each, and 46 997 586 raw SNP variants were subject to multivariable filtering metrics toward a multispecies, genome-wide distributed chip content. Of the 60 904 SNPs on the chip, 59 222 were genotyped and 51 204 were polymorphic across 14 Eucalyptus species, providing a 96% genome-wide coverage with 1 SNP/12-20 kb, and 47 069 SNPs at ≤ 10 kb from 30 444 of the 33 917 genes in the Eucalyptus genome. Given the EUChip60K multi-species genotyping flexibility, we show that both the sample size and taxonomic composition of cluster files impact heterozygous call specificity and sensitivity by benchmarking against 'gold standard' genotypes derived from deeply sequenced individual tree genomes. Thousands of SNPs were shared across species, likely representing ancient variants arisen before the split of these taxa, hinting to a recent eucalypt radiation. We show that the variable SNP filtering constraints allowed coverage of the entire site frequency spectrum, mitigating SNP ascertainment bias. The EUChip60K represents an outstanding tool with which to address population genomics questions in Eucalyptus and to empower genomic selection, GWAS and the broader study of complex trait variation in eucalypts.

Assignment of homoeologs to parental genomes in allopolyploids for species tree inference, with an example from Fumaria (papaveraceae).

There is a rising awareness that species trees are best inferred from multiple loci while taking into account processes affecting individual gene trees, such as substitution model error (failure of the model to account for the complexity of the data) and coalescent stochasticity (presence of incomplete lineage sorting [ILS]). Although most studies have been carried out in the context of dichotomous species trees, these processes operate also in more complex evolutionary histories involving multiple hybridizations and polyploidy. Recently, methods have been developed that accurately handle ILS in allopolyploids, but they are thus far restricted to networks of diploids and tetraploids. We propose a procedure that improves on this limitation by designing a workflow that assigns homoeologs to hypothetical diploid ancestral genomes prior to genome tree construction. Conflicting assignment hypotheses are evaluated against substitution model error and coalescent stochasticity. Incongruence that cannot be explained by stochastic mechanisms needs to be explained by other processes (e.g., homoploid hybridization or paralogy). The data can then be filtered to build multilabeled genome phylogenies using inference methods that can recover species trees, either in the face of substitution model error and coalescent stochasticity alone, or while simultaneously accounting for hybridization. Methods are already available for folding the resulting multilabeled genome phylogeny into a network. We apply the workflow to the reconstruction of the reticulate phylogeny of the plant genus Fumaria (Papaveraceae) with ploidal levels ranging from 2[Formula: see text] to 14[Formula: see text]. We describe the challenges in recovering nuclear NRPB2 homoeologs in high ploidy species while combining in vivo cloning and direct sequencing techniques. Using parametric bootstrapping simulations we assign nuclear homoeologs and chloroplast sequences (four concatenated loci) to their common hypothetical diploid ancestral genomes. As these assignments hinge on effective population size assumptions, we investigate how varying these assumptions impacts the recovered multilabeled genome phylogeny.

Shotgun bisulfite sequencing of the Betula platyphylla genome reveals the tree's DNA methylation patterning.

DNA methylation plays a critical role in the regulation of gene expression. Most studies of DNA methylation have been performed in herbaceous plants, and little is known about the methylation patterns in tree genomes. In the present study, we generated a map of methylated cytosines at single base pair resolution for Betula platyphylla (white birch) by bisulfite sequencing combined with transcriptomics to analyze DNA methylation and its effects on gene expression. We obtained a detailed view of the function of DNA methylation sequence composition and distribution in the genome of B. platyphylla. There are 34,460 genes in the whole genome of birch, and 31,297 genes are methylated. Conservatively, we estimated that 14.29% of genomic cytosines are methylcytosines in birch. Among the methylation sites, the CHH context accounts for 48.86%, and is the largest proportion. Combined transcriptome and methylation analysis showed that the genes with moderate methylation levels had higher expression levels than genes with high and low methylation. In addition, methylated genes are highly enriched for the GO subcategories of binding activities, catalytic activities, cellular processes, response to stimulus and cell death, suggesting that methylation mediates these pathways in birch trees.

Tree Genetics & Genomes introduces Continuous Article Publishing

Tree Genetics & Genomes introduces Continuous Article Publishing

From gene trees to a dated allopolyploid network: insights from the angiosperm genus Viola (Violaceae).

Allopolyploidization accounts for a significant fraction of speciation events in many eukaryotic lineages. However, existing phylogenetic and dating methods require tree-like topologies and are unable to handle the network-like phylogenetic relationships of lineages containing allopolyploids. No explicit framework has so far been established for evaluating competing network topologies, and few attempts have been made to date phylogenetic networks. We used a four-step approach to generate a dated polyploid species network for the cosmopolitan angiosperm genus Viola L. (Violaceae Batch.). The genus contains ca 600 species and both recent (neo-) and more ancient (meso-) polyploid lineages distributed over 16 sections. First, we obtained DNA sequences of three low-copy nuclear genes and one chloroplast region, from 42 species representing all 16 sections. Second, we obtained fossil-calibrated chronograms for each nuclear gene marker. Third, we determined the most parsimonious multilabeled genome tree and its corresponding network, resolved at the section (not the species) level. Reconstructing the “correct” network for a set of polyploids depends on recovering all homoeologs, i.e., all subgenomes, in these polyploids. Assuming the presence of Viola subgenome lineages that were not detected by the nuclear gene phylogenies (“ghost subgenome lineages”) significantly reduced the number of inferred polyploidization events. We identified the most parsimonious network topology from a set of five competing scenarios differing in the interpretation of homoeolog extinctions and lineage sorting, based on (i) fewest possible ghost subgenome lineages, (ii) fewest possible polyploidization events, and (iii) least possible deviation from expected ploidy as inferred from available chromosome counts of the involved polyploid taxa. Finally, we estimated the homoploid and polyploid speciation times of the most parsimonious network. Homoploid speciation times were estimated by coalescent analysis of gene tree node ages. Polyploid speciation times were estimated by comparing branch lengths and speciation rates of lineages with and without ploidy shifts. Our analyses recognize Viola as an old genus (crown age 31 Ma) whose evolutionary history has been profoundly affected by allopolyploidy. Between 16 and 21 allopolyploidizations are necessary to explain the diversification of the 16 major lineages (sections) of Viola, suggesting that allopolyploidy has accounted for a high percentage—between 67% and 88%—of the speciation events at this level. The theoretical and methodological approaches presented here for (i) constructing networks and (ii) dating speciation events within a network, have general applicability for phylogenetic studies of groups where allopolyploidization has occurred. They make explicit use of a hitherto underexplored source of ploidy information from chromosome counts to help resolve phylogenetic cases where incomplete sequence data hampers network inference. Importantly, the coalescent-based method used herein circumvents the assumption of tree-like evolution required by most techniques for dating speciation events.

Subtyping of renal cortical neoplasms in fine needle aspiration biopsies using a decision tree based on genomic alterations detected by fluorescencein situhybridization

ObjectivesTo improve the overall accuracy of diagnosis in needle biopsies of renal masses, especially small renal masses (SRMs), using fluorescence in situ hybridization (FISH), and to develop a renal cortical neoplasm classification decision tree based on genomic alterations detected by FISH.Patients and MethodsEx vivo fine needle aspiration biopsies of 122 resected renal cortical neoplasms were subjected to FISH using a series of seven-probe sets to assess gain or loss of 10 chromosomes and rearrangement of the 11q13 locus. Using specimen (nephrectomy)-histology as the ‘gold standard’, a genomic aberration-based decision tree was generated to classify specimens. The diagnostic potential of the decision tree was assessed by comparing the FISH-based classification and biopsy histology with specimen histology.ResultsOf the 114 biopsies diagnostic by either method, a higher diagnostic yield was achieved by FISH (92 and 96%) than histology alone (82 and 84%) in the 65 biopsies from SRMs (<4 cm) and 49 from larger masses, respectively. An optimized decision tree was constructed based on aberrations detected in eight chromosomes, by which the maximum concordance of classification achieved by FISH was 79%, irrespective of mass size. In SRMs, the overall sensitivity of diagnosis by FISH compared with histopathology was higher for benign oncocytoma, was similar for the chromophobe renal cell carcinoma subtype, and was lower for clear-cell and papillary subtypes. The diagnostic accuracy of classification of needle biopsy specimens (from SRMs) increased from 80% obtained by histology alone to 94% when combining histology and FISH.ConclusionThe present study suggests that a novel FISH assay developed by us has a role to play in assisting in the yield and accuracy of diagnosis of renal cortical neoplasms in needle biopsies in particular, and can help guide the clinical management of patients with SRMs that were non-diagnostic by histology.

CGBayesNets: conditional Gaussian Bayesian network learning and inference with mixed discrete and continuous data.

Bayesian Networks (BN) have been a popular predictive modeling formalism in bioinformatics, but their application in modern genomics has been slowed by an inability to cleanly handle domains with mixed discrete and continuous variables. Existing free BN software packages either discretize continuous variables, which can lead to information loss, or do not include inference routines, which makes prediction with the BN impossible. We present CGBayesNets, a BN package focused around prediction of a clinical phenotype from mixed discrete and continuous variables, which fills these gaps. CGBayesNets implements Bayesian likelihood and inference algorithms for the conditional Gaussian Bayesian network (CGBNs) formalism, one appropriate for predicting an outcome of interest from, e.g., multimodal genomic data. We provide four different network learning algorithms, each making a different tradeoff between computational cost and network likelihood. CGBayesNets provides a full suite of functions for model exploration and verification, including cross validation, bootstrapping, and AUC manipulation. We highlight several results obtained previously with CGBayesNets, including predictive models of wood properties from tree genomics, leukemia subtype classification from mixed genomic data, and robust prediction of intensive care unit mortality outcomes from metabolomic profiles. We also provide detailed example analysis on public metabolomic and gene expression datasets. CGBayesNets is implemented in MATLAB and available as MATLAB source code, under an Open Source license and anonymous download at http://www.cgbayesnets.com.

The genome of Eucalyptus grandis.

Eucalypts are the world's most widely planted hardwood trees. Their outstanding diversity, adaptability and growth have made them a global renewable resource of fibre and energy. We sequenced and assembled >94% of the 640-megabase genome of Eucalyptus grandis. Of 36,376 predicted protein-coding genes, 34% occur in tandem duplications, the largest proportion thus far in plant genomes. Eucalyptus also shows the highest diversity of genes for specialized metabolites such as terpenes that act as chemical defence and provide unique pharmaceutical oils. Genome sequencing of the E. grandis sister species E. globulus and a set of inbred E. grandis tree genomes reveals dynamic genome evolution and hotspots of inbreeding depression. The E. grandis genome is the first reference for the eudicot order Myrtales and is placed here sister to the eurosids. This resource expands our understanding of the unique biology of large woody perennials and provides a powerful tool to accelerate comparative biology, breeding and biotechnology.

Characterization of rubber tree microRNA in phytohormone response using large genomic DNA libraries, promoter sequence and gene expression analysis.

The para rubber tree is the most widely cultivated tree species for producing natural rubber (NR) latex. Unfortunately, rubber tree characteristics such as a long life cycle, heterozygous genetic backgrounds, and poorly understood genetic profiles are the obstacles to breeding new rubber tree varieties, such as those with improved NR yields. Recent evidence has revealed the potential importance of controlling microRNA (miRNA) decay in some aspects of NR regulation. To gain a better understanding of miRNAs and their relationship with rubber tree gene regulation networks, large genomic DNA insert-containing libraries were generated to complement the incomplete draft genome sequence and applied as a new powerful tool to predict a function of interested genes. Bacterial artificial chromosome and fosmid libraries, containing a total of 120,576 clones with an average insert size of 43.35kb, provided approximately 2.42 haploid genome equivalents of coverage based on the estimated 2.15gb rubber tree genome. Based on these library sequences, the precursors of 1 member of rubber tree-specific miRNAs and 12 members of conserved miRNAs were successfully identified. A panel of miRNAs was characterized for phytohormone response by precisely identifying phytohormone-responsive motifs in their promoter sequences. Furthermore, the quantitative real-time PCR on ethylene stimulation of rubber trees was performed to demonstrate that the miR2118, miR159, miR164 and miR166 are responsive to ethylene, thus confirmed the prediction by genomic DNA analysis. The cis-regulatory elements identified in the promoter regions of these miRNA genes help augment our understanding of miRNA gene regulation and provide a foundation for further investigation of the regulation of rubber tree miRNAs.

Genome-wide identification and characterization of WRKY gene family in Hevea brasiliensis

WRKY proteins constitute a large family of transcription factors. In this study, we identified 81 WRKY genes (named HbWRKY1 to HbWRKY81) in the latest rubber tree genome. Tissue-specific expression profiles showed that 74 HbWRKYs were expressed in at least one of the tissues and the other 7 genes showed very low expression in all tissues tested, which suggested that HbWRKYs took part in many cellular processes. The responses of 20 selected HbWRKYs to jasmonic acid (JA) and ethylene (ET) were analyzed in the latex. 17 HbWRKYs responded to at least one treatment, which included 15 HbWRKYs responding to JA treatment, 15 HbWRKYs to ET, which suggested that these HbWRKYs were regulated by JA and ET. We also observed that HbWRKY3, 14, and 55 bind HbSRPP promoter and activate the transcription in yeast. This study suggests that HbWRKY proteins maybe involved in the transcriptional regulation of nature rubber biosynthesis.

Zero tolerance for healthcare-associated MRSA bacteraemia: is it realistic?

BackgroundThe term ‘zero tolerance’ has recently been applied to healthcare-associated infections, implying that such events are always preventable. This may not be the case for healthcare-associated infections such as methicillin-resistant Staphylococcus aureus (MRSA) bacteraemia.MethodsWe combined information from an epidemiological investigation and bacterial whole-genome sequencing to evaluate a cluster of five MRSA bacteraemia episodes in four patients in a specialist hepatology unit.ResultsThe five MRSA bacteraemia isolates were highly related by multilocus sequence type (ST) (four isolates were ST22 and one isolate was a single-locus variant, ST2046). Whole-genome sequencing demonstrated unequivocally that the bacteraemia cases were unrelated. Placing the MRSA bacteraemia isolates within a local and global phylogenetic tree of MRSA ST22 genomes demonstrated that the five bacteraemia isolates were highly diverse. This was consistent with the acquisition and importation of MRSA from the wider referral network. Analysis of MRSA carriage and disease in patients within the hepatology service demonstrated a higher risk of both initial MRSA acquisition compared with the nephrology service and a higher risk of progression from MRSA carriage to bacteraemia, compared with patients in nephrology or geriatric services. A root cause analysis failed to reveal any mechanism by which three of five MRSA bacteraemia episodes could have been prevented.ConclusionsThis study illustrates the complex nature of MRSA carriage and bacteraemia in patients in a specialized hepatology unit. Despite numerous ongoing interventions to prevent MRSA bacteraemia in healthcare settings, these are unlikely to result in a zero incidence in referral centres that treat highly complex patients.

Identification and characterization of the 14-3-3 gene family in Hevea brasiliensis

The 14-3-3 proteins are a family of conserved phospho-specific binding proteins involved in diverse physiological processes. Although the genome-wide analysis of this family has been carried out in certain plant species, little is known about 14-3-3 protein genes in rubber tree (Hevea brasiliensis). In this study, we identified 10 14-3-3 protein genes (designated as HbGF14a to HbGF14j) in the latest rubber tree genome. A phylogenetic tree was constructed and found to demonstrate that HbGF14s can be divided into two major groups. Tissue-specific expression profiles showed that 10 HbGF14 were expressed in at least one of the tissues, which suggested that HbGF14s participated in numerous cellular processes. The 10 HbGF14s responded to jasmonic acid (JA) and ethylene (ET) treatment, which suggested that these HbGF14s were involved in response to JA and ET signaling. The target of HbGF14c protein was related to small rubber particle protein, a major rubber particle protein that is involved in rubber biosynthesis. These findings suggested that 14-3-3 proteins may be involved in the regulation of natural rubber biosynthesis.

&lt;i&gt;In Silico&lt;/i&gt; Analysis of Cellulose Synthase Gene (NcCesA1) in Developing Xylem Tissues of &lt;i&gt;Neolamarckia Cadamba&lt;/i&gt;

This study reported the isolation and in silico characterization of full-length cellulose synthase (CesA) cDNA from Neolamarckia cadamba, an important tropical plantation tree species. CesA is a member of processive glycosyltransferases that involved in cellulose biosynthesis of plants. CesA acts as a central catalyst in the generation of plant cell wall biomass or cellulose. It also plays an important role in regulating wood formation. The hypothetical full-length CesA cDNA (NcCesA1; JX134621) was assembled by contig mapping approach using the CesA cDNA sequences from NcdbEST and the amplified 5�-and 3�-RACE PCR sequences. The NcCesA1 cDNA has a length of 3,472 bp with 3,126 bp open reading frame encoding a 1,042 amino acid sequence. The predicted NcCesA1 protein contained N-terminal cysteine rich zinc binding domain, 7 putative Transmembrane Helices (TMH), 4 U-motifs that contain a signature D, D, D, QxxRW motif, an alternating Conserved Region (CR-P) and 2 Hypervariable Regions (HVR). These entire shared domain structures suggest the functional role of NcCesA1 is involved in glycosyltransferation of the secondary cell wall cellulose biosynthesis of N. cadamba. Sequence comparison also revealed the high similarity (90%) among NcCesA1 and PtrCesA2 of Populus tremuloides. This further implies the involvement of NcCesA1 that catalyzes the cellulose biosynthesis of secondary cell wall rather than primary cell wall. This full-length NcCesA1 cDNA can serve as good candidate gene in association genetics study which leads to Gene-Assisted Selection (GAS) in the N. cadamba tree breeding programme. Furthermore, the isolation of new CesA genes from tropical tree genomes is essential for enhancing knowledge of cellulose biosynthesis in trees that has many fundamental and commercial implications.

The Genome Sequence of ‘Mycobacterium massiliense’ Strain CIP 108297 Suggests the Independent Taxonomic Status of the Mycobacterium abscessus Complex at the Subspecies Level

Members of the Mycobacterium abscessus complex are rapidly growing mycobacteria that are emerging as human pathogens. The M. abscessus complex was previously composed of three species, namely M. abscessus sensu stricto, ‘M. massiliense’, and ‘M. bolletii’. In 2011, ‘M. massiliense’ and ‘M. bolletii’ were united and reclassified as a single subspecies within M. abscessus: M. abscessus subsp. bolletii. However, the placement of ‘M. massiliense’ within the boundary of M. abscessus subsp. bolletii remains highly controversial with regard to clinical aspects. In this study, we revisited the taxonomic status of members of the M. abscessus complex based on comparative analysis of the whole-genome sequences of 53 strains. The genome sequence of the previous type strain of ‘Mycobacterium massiliense’ (CIP 108297) was determined using next-generation sequencing. The genome tree based on average nucleotide identity (ANI) values supported the differentiation of ‘M. bolletii’ and ‘M. massiliense’ at the subspecies level. The genome tree also clearly illustrated that ‘M. bolletii’ and ‘M. massiliense’ form a distinct phylogenetic clade within the radiation of the M. abscessus complex. The genomic distances observed in this study suggest that the current M. abscessus subsp. bolletii taxon should be divided into two subspecies, M. abscessus subsp. massiliense subsp. nov. and M. abscessus subsp. bolletii, to correspondingly accommodate the previously known ‘M. massiliense’ and ‘M. bolletii’ strains.

An overview of forestry journals in the period 2006–2010 as basis for ascertaining research trends

Articles published between January 1, 2006 and December 31, 2010 in 42 forestry journals (N = 16,258) were collected and, depending on their content and key words, classified in one of 22 sub-disciplines. Among the forestry sub-disciplines, the following are currently dominant: Mensuration and inventories, Forest management, Plant ecophysiology and Wood science. PCA ordination was used to visualize grouping tendencies and data separation. For each component, a number of characteristics contributed to the total variation, and significant importance was attached to those with the highest loading factors. The first component included Mensuration and inventories, Plant ecophysiology, Vegetation ecology and Forest management, as the highest loading factors. The second components comprised Sociological aspects, Plant ecophysiology, Wood science and Forest management. The most pronounced increase trend over the five-year period is noted for Genetics and breeding, Vegetation ecology, Fuels and energy, while the most pronounced decrease trend is visible in Forest health, Forest fire, Sociological aspects and Forest products. PCA suggests the existence of three groups of journals: the first group comprises Forest Ecology and Management and Canadian Journal of Forest Research, the dominating two, the second group comprises Annals of Forest Science, Plant Ecology, Tree Physiology and Trees-Structure and Function, while the rest of the journals belong to the third group. The Canadian Journal of Forest Research is the most diversified, while Tree Genetics and Genomes, Silvae Genetica and Tree-ring Research are narrowly specialized.

Construction of a Phylogenetic Tree of Photosynthetic Prokaryotes Based on Average Similarities of Whole Genome Sequences

Phylogenetic trees have been constructed for a wide range of organisms using gene sequence information, especially through the identification of orthologous genes that have been vertically inherited. The number of available complete genome sequences is rapidly increasing, and many tools for construction of genome trees based on whole genome sequences have been proposed. However, development of a reasonable method of using complete genome sequences for construction of phylogenetic trees has not been established. We have developed a method for construction of phylogenetic trees based on the average sequence similarities of whole genome sequences. We used this method to examine the phylogeny of 115 photosynthetic prokaryotes, i.e., cyanobacteria, Chlorobi, proteobacteria, Chloroflexi, Firmicutes and nonphotosynthetic organisms including Archaea. Although the bootstrap values for the branching order of phyla were low, probably due to lateral gene transfer and saturated mutation, the obtained tree was largely consistent with the previously reported phylogenetic trees, indicating that this method is a robust alternative to traditional phylogenetic methods.

Alignment-Free Genome Tree Inference by Learning Group-Specific Distance Metrics

Understanding the evolutionary relationships between organisms is vital for their in-depth study. Gene-based methods are often used to infer such relationships, which are not without drawbacks. One can now attempt to use genome-scale information, because of the ever increasing number of genomes available. This opportunity also presents a challenge in terms of computational efficiency. Two fundamentally different methods are often employed for sequence comparisons, namely alignment-based and alignment-free methods. Alignment-free methods rely on the genome signature concept and provide a computationally efficient way that is also applicable to nonhomologous sequences. The genome signature contains evolutionary signal as it is more similar for closely related organisms than for distantly related ones. We used genome-scale sequence information to infer taxonomic distances between organisms without additional information such as gene annotations. We propose a method to improve genome tree inference by learning specific distance metrics over the genome signature for groups of organisms with similar phylogenetic, genomic, or ecological properties. Specifically, our method learns a Mahalanobis metric for a set of genomes and a reference taxonomy to guide the learning process. By applying this method to more than a thousand prokaryotic genomes, we showed that, indeed, better distance metrics could be learned for most of the 18 groups of organisms tested here. Once a group-specific metric is available, it can be used to estimate the taxonomic distances for other sequenced organisms from the group. This study also presents a large scale comparison between 10 methods—9 alignment-free and 1 alignment-based.

Sprucing up forest tree genomics

Sprucing up forest tree genomics

Open access to tree genomes: the path to a better forest

An open-access culture and a well-developed comparative-genomics infrastructure must be developed in forest trees to derive the full potential of genome sequencing in this diverse group of plants that are the dominant species in much of the earth's terrestrial ecosystems.

Putting the landscape into the genomics of trees: approaches for understanding local adaptation and population responses to changing climate

The Forest ecosystem genomics Research: supporTing Transatlantic Cooperation project (FoResTTraC, http://www.foresttrac.eu/ ) sponsored a workshop in August 2010 to evaluate the potential for using a landscape genomics approach for studying plant adaptation to the environment and the potential of local populations for coping with changing climate. This paper summarizes our discussions and articulates a vision of how we believe forest trees offer an unparalleled opportunity to address fundamental biological questions, as well as how the application of landscape genomic methods complement to traditional forest genetic approaches that provide critical information needed for natural resource management. In this paper, we will cover four topics. First, we begin by defining landscape genomics and briefly reviewing the unique situation for tree species in the application of this approach toward understanding plant adaptation to the environment. Second, we review traditional approaches in forest genetics for studying local adaptation and identifying loci underlying locally adapted phenotypes. Third, we present existing and emerging methods available for landscape genomic analyses. Finally, we briefly touch on how these approaches can aid in understanding practical topics such as management of tree populations facing climate change.