The ELIXIR Biodiversity Community: Understanding short- and long-term changes in biodiversity.

Biodiversity loss is now recognised as one of the major challenges for humankind to address over the next few decades. Unless major actions are taken, the sixth mass extinction will lead to catastrophic effects on the Earth's biosphere and human health and well-being. ELIXIR can help address the technical challenges of biodiversity science, through leveraging its suite of services and expertise to enable data management and analysis activities that enhance our understanding of life on Earth and facilitate biodiversity preservation and restoration. This white paper, prepared by the ELIXIR Biodiversity Community, summarises the current status and responses, and presents a set of plans, both technical and community-oriented, that should both enhance how ELIXIR Services are applied in the biodiversity field and how ELIXIR builds connections across the many other infrastructures active in this area. We discuss the areas of highest priority, how they can be implemented in cooperation with the ELIXIR Platforms, and their connections to existing ELIXIR Communities and international consortia. The article provides a preliminary blueprint for a Biodiversity Community in ELIXIR and is an appeal to identify and involve new stakeholders.

A widespread inversion polymorphism conserved among Saccharomyces species is caused by recurrent homogenization of a sporulation gene family.

Saccharomyces genomes are highly collinear and show relatively little structural variation, both within and between species of this yeast genus. We investigated the only common inversion polymorphism known in S. cerevisiae, which affects a 24-kb 'flip/flop' region containing 15 genes near the centromere of chromosome XIV. The region exists in two orientations, called reference (REF) and inverted (INV). Meiotic recombination in this region is suppressed in crosses between REF and INV orientation strains such as the BY x RM cross. We find that the inversion polymorphism is at least 17 million years old because it is conserved across the genus Saccharomyces. However, the REF and INV isomers are not ancient alleles but are continually being re-created by re-inversion of the region within each species. Inversion occurs due to continual homogenization of two almost identical 4-kb sequences that form an inverted repeat (IR) at the ends of the flip/flop region. The IR consists of two pairs of genes that are specifically and strongly expressed during the late stages of sporulation. We show that one of these gene pairs, YNL018C/YNL034W, codes for a protein that is essential for spore formation. YNL018C and YNL034W are the founder members of a gene family, Centroid, whose members in other Saccharomycetaceae species evolve fast, duplicate frequently, and are preferentially located close to centromeres. We tested the hypothesis that Centroid genes are a meiotic drive system, but found no support for this idea.

Contradictory Phylogenetic Signals in the Laurasiatheria Anomaly Zone.

Relationships among laurasiatherian clades represent one of the most highly disputed topics in mammalian phylogeny. In this study, we attempt to disentangle laurasiatherian interordinal relationships using two independent genome-level approaches: (1) quantifying retrotransposon presence/absence patterns, and (2) comparisons of exon datasets at the levels of nucleotides and amino acids. The two approaches revealed contradictory phylogenetic signals, possibly due to a high level of ancestral incomplete lineage sorting. The positions of Eulipotyphla and Chiroptera as the first and second earliest divergences were consistent across the approaches. However, the phylogenetic relationships of Perissodactyla, Cetartiodactyla, and Ferae, were contradictory. While retrotransposon insertion analyses suggest a clade with Cetartiodactyla and Ferae, the exon dataset favoured Cetartiodactyla and Perissodactyla. Future analyses of hitherto unsampled laurasiatherian lineages and synergistic analyses of retrotransposon insertions, exon and conserved intron/intergenic sequences might unravel the conflicting patterns of relationships in this major mammalian clade.

Genetic and lipidomic analyses suggest that Nostoc punctiforme, a plant-symbiotic cyanobacterium, does not produce sphingolipids.

Sphingolipids, a class of amino-alcohol-based lipids, are well characterized in eukaryotes and in some anaerobic bacteria. However, the only sphingolipids so far identified in cyanobacteria are two ceramides (i.e., an acetylsphingomyelin and a cerebroside), both based on unbranched, long-chain base (LCB) sphingolipids in Scytonema julianum and Moorea producens , respectively. The first step in de novo sphingolipid biosynthesis is the condensation of l-serine with palmitoyl-CoA to produce 3-keto-diyhydrosphingosine (KDS). This reaction is catalyzed by serine palmitoyltransferase (SPT), which belongs to a small family of pyridoxal phosphate-dependent α-oxoamine synthase (AOS) enzymes. Based on sequence similarity to molecularly characterized bacterial SPT peptides, we identified a putative SPT (Npun_R3567) from the model nitrogen-fixing, plant-symbiotic cyanobacterium, Nostoc punctiforme strain PCC 73102 (ATCC 29133). Gene expression analysis revealed that Npun_R3567 is induced during late-stage diazotrophic growth in N. punctiforme . However, Npun_R3567 could not produce the SPT reaction product, 3-keto-diyhydrosphingosine (KDS), when heterologously expressed in Escherichia coli . This agreed with a sphingolipidomic analysis of N. punctiforme cells, which revealed that no LCBs or ceramides were present. To gain a better understanding of Npun_R3567, we inferred the phylogenetic position of Npun_R3567 relative to other bacterial AOS peptides. Rather than clustering with other bacterial SPTs, Npun_R3567 and the other cyanobacterial BioF homologues formed a separate, monophyletic group. Given that N. punctiforme does not appear to possess any other gene encoding an AOS enzyme, it is altogether unlikely that N. punctiforme is capable of synthesizing sphingolipids. In the context of cross-kingdom symbiosis signalling in which sphingolipids are emerging as important regulators, it appears unlikely that sphingolipids from N. punctiforme play a regulatory role during its symbiotic association with plants.

A Likelihood-Ratio Test for Lumpability of Phylogenetic Data: Is the Markovian Property of an Evolutionary Process Retained in Recoded DNA?

In molecular phylogenetics, it is typically assumed that the evolutionary process for DNA can be approximated by independent and identically distributed Markovian processes at the variable sites and that these processes diverge over the edges of a rooted bifurcating tree. Sometimes the nucleotides are transformed from a 4-state alphabet to a 3- or 2-state alphabet by a procedure that is called recoding, lumping, or grouping of states. Here, we introduce a likelihood-ratio test for lumpability for DNA that has diverged under different Markovian conditions, which assesses the assumption that the Markovian property of the evolutionary process over each edge is retained after recoding of the nucleotides. The test is derived and validated numerically on simulated data. To demonstrate the insights that can be gained by using the test, we assessed two published data sets, one of mitochondrial DNA from a phylogenetic study of the ratites and the other of nuclear DNA from a phylogenetic study of yeast. Our analysis of these data sets revealed that recoding of the DNA eliminated some of the compositional heterogeneity detected over the sequences. However, the Markovian property of the original evolutionary process was not retained by the recoding, leading to some significant distortions of edge lengths in reconstructed trees.[Evolutionary processes; likelihood-ratio test; lumpability; Markovian processes; Markov models; phylogeny; recoding of nucleotides.].

A globally diverse reference alignment and panel for imputation of mitochondrial DNA variants.

BACKGROUND: Variation in mitochondrial DNA (mtDNA) identified by genotyping microarrays or by sequencing only the hypervariable regions of the genome may be insufficient to reliably assign mitochondrial genomes to phylogenetic lineages or haplogroups. This lack of resolution can limit functional and clinical interpretation of a substantial body of existing mtDNA data. To address this limitation, we developed and evaluated a large, curated reference alignment of complete mtDNA sequences as part of a pipeline for imputing missing mtDNA single nucleotide variants (mtSNVs). We call our reference alignment and pipeline MitoImpute. RESULTS: We aligned the sequences of 36,960 complete human mitochondrial genomes downloaded from GenBank, filtered and controlled for quality. These sequences were reformatted for use in imputation software, IMPUTE2. We assessed the imputation accuracy of MitoImpute by measuring haplogroup and genotype concordance in data from the 1000 Genomes Project and the Alzheimer's Disease Neuroimaging Initiative (ADNI). The mean improvement of haplogroup assignment in the 1000 Genomes samples was 42.7% (Matthew's correlation coefficient = 0.64). In the ADNI cohort, we imputed missing single nucleotide variants. CONCLUSION: These results show that our reference alignment and panel can be used to impute missing mtSNVs in existing data obtained from using microarrays, thereby broadening the scope of functional and clinical investigation of mtDNA. This improvement may be particularly useful in studies where participants have been recruited over time and mtDNA data obtained using different methods, enabling better integration of early data collected using less accurate methods with more recent sequence data.

Analysis of Genomic Sequence Data Reveals the Origin and Evolutionary Separation of Hawaiian Hoary Bat Populations.

We examine the genetic history and population status of Hawaiian hoary bats (Lasiurus semotus), the most isolated bats on Earth, and their relationship to northern hoary bats (Lasiurus cinereus), through whole-genome analysis of single-nucleotide polymorphisms mapped to a de novo-assembled reference genome. Profiles of genomic diversity and divergence indicate that Hawaiian hoary bats are distinct from northern hoary bats, and form a monophyletic group, indicating a single ancestral colonization event 1.34 Ma, followed by substantial divergence between islands beginning 0.51 Ma. Phylogenetic analysis indicates Maui is central to the radiation across the archipelago, with the southward expansion to Hawai'i and westward to O'ahu and Kaua'i. Because this endangered species is of conservation concern, a clearer understanding of the population genetic structure of this bat in the Hawaiian Islands is of timely importance.

Six reference-quality genomes reveal evolution of bat adaptations.

Bats possess extraordinary adaptations, including flight, echolocation, extreme longevity and unique immunity. High-quality genomes are crucial for understanding the molecular basis and evolution of these traits. Here we incorporated long-read sequencing and state-of-the-art scaffolding protocols1 to generate, to our knowledge, the first reference-quality genomes of six bat species (Rhinolophus ferrumequinum, Rousettus aegyptiacus, Phyllostomus discolor, Myotis myotis, Pipistrellus kuhlii and Molossus molossus). We integrated gene projections from our 'Tool to infer Orthologs from Genome Alignments' (TOGA) software with de novo and homology gene predictions as well as short- and long-read transcriptomics to generate highly complete gene annotations. To resolve the phylogenetic position of bats within Laurasiatheria, we applied several phylogenetic methods to comprehensive sets of orthologous protein-coding and noncoding regions of the genome, and identified a basal origin for bats within Scrotifera. Our genome-wide screens revealed positive selection on hearing-related genes in the ancestral branch of bats, which is indicative of laryngeal echolocation being an ancestral trait in this clade. We found selection and loss of immunity-related genes (including pro-inflammatory NF-κB regulators) and expansions of anti-viral APOBEC3 genes, which highlights molecular mechanisms that may contribute to the exceptional immunity of bats. Genomic integrations of diverse viruses provide a genomic record of historical tolerance to viral infection in bats. Finally, we found and experimentally validated bat-specific variation in microRNAs, which may regulate bat-specific gene-expression programs. Our reference-quality bat genomes provide the resources required to uncover and validate the genomic basis of adaptations of bats, and stimulate new avenues of research that are directly relevant to human health and disease1.

A minimum reporting standard for multiple sequence alignments.

Multiple sequence alignments (MSAs) play a pivotal role in studies of molecular sequence data, but nobody has developed a minimum reporting standard (MRS) to quantify the completeness of MSAs in terms of completely specified nucleotides or amino acids. We present an MRS that relies on four simple completeness metrics. The metrics are implemented in AliStat, a program developed to support the MRS. A survey of published MSAs illustrates the benefits and unprecedented transparency offered by the MRS.

A new phylogenetic protocol: dealing with model misspecification and confirmation bias in molecular phylogenetics.

Molecular phylogenetics plays a key role in comparative genomics and has increasingly significant impacts on science, industry, government, public health and society. In this paper, we posit that the current phylogenetic protocol is missing two critical steps, and that their absence allows model misspecification and confirmation bias to unduly influence phylogenetic estimates. Based on the potential offered by well-established but under-used procedures, such as assessment of phylogenetic assumptions and tests of goodness of fit, we introduce a new phylogenetic protocol that will reduce confirmation bias and increase the accuracy of phylogenetic estimates.

GHOST: Recovering Historical Signal from Heterotachously Evolved Sequence Alignments.

Molecular sequence data that have evolved under the influence of heterotachous evolutionary processes are known to mislead phylogenetic inference. We introduce the General Heterogeneous evolution On a Single Topology (GHOST) model of sequence evolution, implemented under a maximum-likelihood framework in the phylogenetic program IQ-TREE (http://www.iqtree.org). Simulations show that using the GHOST model, IQ-TREE can accurately recover the tree topology, branch lengths, and substitution model parameters from heterotachously evolved sequences. We investigate the performance of the GHOST model on empirical data by sampling phylogenomic alignments of varying lengths from a plastome alignment. We then carry out inference under the GHOST model on a phylogenomic data set composed of 248 genes from 16 taxa, where we find the GHOST model concurs with the currently accepted view, placing turtles as a sister lineage of archosaurs, in contrast to results obtained using traditional variable rates-across-sites models. Finally, we apply the model to a data set composed of a sodium channel gene of 11 fish taxa, finding that the GHOST model is able to elucidate a subtle component of the historical signal, linked to the previously established convergent evolution of the electric organ in two geographically distinct lineages of electric fish. We compare inference under the GHOST model to partitioning by codon position and show that, owing to the minimization of model constraints, the GHOST model offers unique biological insights when applied to empirical data.

Sensitivity and specificity of information criteria.

Information criteria (ICs) based on penalized likelihood, such as Akaike's information criterion (AIC), the Bayesian information criterion (BIC) and sample-size-adjusted versions of them, are widely used for model selection in health and biological research. However, different criteria sometimes support different models, leading to discussions about which is the most trustworthy. Some researchers and fields of study habitually use one or the other, often without a clearly stated justification. They may not realize that the criteria may disagree. Others try to compare models using multiple criteria but encounter ambiguity when different criteria lead to substantively different answers, leading to questions about which criterion is best. In this paper we present an alternative perspective on these criteria that can help in interpreting their practical implications. Specifically, in some cases the comparison of two models using ICs can be viewed as equivalent to a likelihood ratio test, with the different criteria representing different alpha levels and BIC being a more conservative test than AIC. This perspective may lead to insights about how to interpret the ICs in more complex situations. For example, AIC or BIC could be preferable, depending on the relative importance one assigns to sensitivity versus specificity. Understanding the differences and similarities among the ICs can make it easier to compare their results and to use them to make informed decisions.

Genome-wide analysis of MIKC-type MADS-box genes in wheat: pervasive duplications, functional conservation and putative neofunctionalization.

Wheat (Triticum aestivum) is one of the most important crops worldwide. Given a growing global population coupled with increasingly challenging cultivation conditions, facilitating wheat breeding by fine-tuning important traits is of great importance. MADS-box genes are prime candidates for this, as they are involved in virtually all aspects of plant development. Here, we present a detailed overview of phylogeny and expression of 201 wheat MIKC-type MADS-box genes. Homoeolog retention is significantly above the average genome-wide retention rate for wheat genes, indicating that many MIKC-type homoeologs are functionally important and not redundant. Gene expression is generally in agreement with the expected subfamily-specific expression pattern, indicating broad conservation of function of MIKC-type genes during wheat evolution. We also found extensive expansion of some MIKC-type subfamilies, especially those potentially involved in adaptation to different environmental conditions like flowering time genes. Duplications are especially prominent in distal telomeric regions. A number of MIKC-type genes show novel expression patterns and respond, for example, to biotic stress, pointing towards neofunctionalization. We speculate that conserved, duplicated and neofunctionalized MIKC-type genes may have played an important role in the adaptation of wheat to a diversity of conditions, hence contributing to the importance of wheat as a global staple food.

Serpins: Genome-Wide Characterisation and Expression Analysis of the Serine Protease Inhibitor Family in Triticum aestivum.

The serine protease inhibitor (serpin) gene family is the largest family of protease inhibitors. Serine protease inhibitors have an active, but under-characterized, role in grain development and defense against pathogen attack in cereal crops. By exploiting publicly available genomic, transcriptomic and proteomic data for wheat (Triticum aestivum), we have identified and annotated the entire 'serpinome' of wheat and constructed a high-quality and robust phylogenetic tree of the gene family, identifying paralogous and homeologous clades from the hexaploid wheat genome, including the Serpin-Z group that have been well characterized in barley. Using publicly available RNAseq data (http://www.wheat-expression.com/), expression profiles of the wheat serpins were explored across a variety of tissues from the developing grain, spikelet and spike. We show that the SERPIN-Z clade, among others, are highly expressed during grain development, and that there is homeologous and paralogous functional redundancy in this gene family. Further to their role in grain development, serpins play an important but under-explored role in response to fungal pathogens. Using 13 RNAseq datasets of wheat tissues infected by fungal pathogens, we identified 37 serpins with a significant disease response. The majority of the disease-responsive serpins were upregulated by Fusarium graminearum, a destructive fungal pathogen that attacks the spike and developing grain of wheat. As serpins are ubiquitous in wheat grain, the genes encoding serpins may be linked to grain development, with their disease response a result of pleiotropy.

Molecular phylogeny reveals food plasticity in the evolution of true ladybird beetles (Coleoptera: Coccinellidae: Coccinellini).

BACKGROUND: The tribe Coccinellini is a group of relatively large ladybird beetles that exhibits remarkable morphological and biological diversity. Many species are aphidophagous, feeding as larvae and adults on aphids, but some species also feed on other hemipterous insects (i.e., heteropterans, psyllids, whiteflies), beetle and moth larvae, pollen, fungal spores, and even plant tissue. Several species are biological control agents or widespread invasive species (e.g., Harmonia axyridis (Pallas)). Despite the ecological importance of this tribe, relatively little is known about the phylogenetic relationships within it. The generic concepts within the tribe Coccinellini are unstable and do not reflect a natural classification, being largely based on regional revisions. This impedes the phylogenetic study of important traits of Coccinellidae at a global scale (e.g. the evolution of food preferences and biogeography). RESULTS: We present the most comprehensive phylogenetic analysis of Coccinellini to date, based on three nuclear and one mitochondrial gene sequences of 38 taxa, which represent all major Coccinellini lineages. The phylogenetic reconstruction supports the monophyly of Coccinellini and its sister group relationship to Chilocorini. Within Coccinellini, three major clades were recovered that do not correspond to any previously recognised divisions, questioning the traditional differentiation between Halyziini, Discotomini, Tytthaspidini, and Singhikaliini. Ancestral state reconstructions of food preferences and morphological characters support the idea of aphidophagy being the ancestral state in Coccinellini. This indicates a transition from putative obligate scale feeders, as seen in the closely related Chilocorini, to more agile general predators. CONCLUSIONS: Our results suggest that the classification of Coccinellini has been misled by convergence in morphological traits. The evolutionary history of Coccinellini has been very dynamic in respect to changes in host preferences, involving multiple independent host switches from different insect orders to fungal spores and plants tissues. General predation on ephemeral aphids might have created an opportunity to easily adapt to mixed or specialised diets (e.g. obligate mycophagy, herbivory, predation on various hemipteroids or larvae of leaf beetles (Chrysomelidae)). The generally long-lived adults of Coccinellini can consume pollen and floral nectars, thereby surviving periods of low prey frequency. This capacity might have played a central role in the diversification history of Coccinellini.

ModelFinder: fast model selection for accurate phylogenetic estimates.

Model-based molecular phylogenetics plays an important role in comparisons of genomic data, and model selection is a key step in all such analyses. We present ModelFinder, a fast model-selection method that greatly improves the accuracy of phylogenetic estimates by incorporating a model of rate heterogeneity across sites not previously considered in this context and by allowing concurrent searches of model space and tree space.

Mitochondrial DNA and trade data support multiple origins of Helicoverpa armigera (Lepidoptera, Noctuidae) in Brazil.

The Old World bollworm Helicoverpa armigera is now established in Brazil but efforts to identify incursion origin(s) and pathway(s) have met with limited success due to the patchiness of available data. Using international agricultural/horticultural commodity trade data and mitochondrial DNA (mtDNA) cytochrome oxidase I (COI) and cytochrome b (Cyt b) gene markers, we inferred the origins and incursion pathways into Brazil. We detected 20 mtDNA haplotypes from six Brazilian states, eight of which were new to our 97 global COI-Cyt b haplotype database. Direct sequence matches indicated five Brazilian haplotypes had Asian, African, and European origins. We identified 45 parsimoniously informative sites and multiple substitutions per site within the concatenated (945 bp) nucleotide dataset, implying that probabilistic phylogenetic analysis methods are needed. High diversity and signatures of uniquely shared haplotypes with diverse localities combined with the trade data suggested multiple incursions and introduction origins in Brazil. Increasing agricultural/horticultural trade activities between the Old and New Worlds represents a significant biosecurity risk factor. Identifying pest origins will enable resistance profiling that reflects countries of origin to be included when developing a resistance management strategy, while identifying incursion pathways will improve biosecurity protocols and risk analysis at biosecurity hotspots including national ports.

Identifying Optimal Models of Evolution.

Most phylogenetic methods are model-based and depend on models of evolution designed to approximate the evolutionary processes. Several methods have been developed to identify suitable models of evolution for phylogenetic analysis of alignments of nucleotide or amino acid sequences and some of these methods are now firmly embedded in the phylogenetic protocol. However, in a disturbingly large number of cases, it appears that these models were used without acknowledgement of their inherent shortcomings. In this chapter, we discuss the problem of model selection and show how some of the inherent shortcomings may be identified and overcome.

Are feeding preferences and insecticide resistance associated with the size of detoxifying enzyme families in insect herbivores?

The size of gene families associated with xenobiotic detoxification in insects may be associated with the complexity of their diets and their propensities to develop insecticide resistance. We test these hypotheses by collating the annotations of cytochrome P450, carboxyl/cholinesterase and glutathione S-transferase genes in 65 insect species with data on their host use and history of insecticide resistance. We find 2-4 fold variation across the species in the numbers of these genes and, in some orders, especially the Hymenoptera, there is a clear relationship between the numbers of genes and feeding preferences. However in other orders, in particular the Lepidoptera, no such relationship is apparent. The size of these three gene families also tend to correlate with insecticide resistance propensity but this may not be an independent effect because species with broader host ranges are more likely to be pests that are heavily sprayed with insecticides.

Phylogenomics resolves the timing and pattern of insect evolution.

Insects are the most speciose group of animals, but the phylogenetic relationships of many major lineages remain unresolved. We inferred the phylogeny of insects from 1478 protein-coding genes. Phylogenomic analyses of nucleotide and amino acid sequences, with site-specific nucleotide or domain-specific amino acid substitution models, produced statistically robust and congruent results resolving previously controversial phylogenetic relations hips. We dated the origin of insects to the Early Ordovician [~479 million years ago (Ma)], of insect flight to the Early Devonian (~406 Ma), of major extant lineages to the Mississippian (~345 Ma), and the major diversification of holometabolous insects to the Early Cretaceous. Our phylogenomic study provides a comprehensive reliable scaffold for future comparative analyses of evolutionary innovations among insects.