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 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.

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.].

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.

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.

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.

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.

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.

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.

Mixture models of nucleotide sequence evolution that account for heterogeneity in the substitution process across sites and across lineages.

Molecular phylogenetic studies of homologous sequences of nucleotides often assume that the underlying evolutionary process was globally stationary, reversible, and homogeneous (SRH), and that a model of evolution with one or more site-specific and time-reversible rate matrices (e.g., the GTR rate matrix) is enough to accurately model the evolution of data over the whole tree. However, an increasing body of data suggests that evolution under these conditions is an exception, rather than the norm. To address this issue, several non-SRH models of molecular evolution have been proposed, but they either ignore heterogeneity in the substitution process across sites (HAS) or assume it can be modeled accurately using the distribution. As an alternative to these models of evolution, we introduce a family of mixture models that approximate HAS without the assumption of an underlying predefined statistical distribution. This family of mixture models is combined with non-SRH models of evolution that account for heterogeneity in the substitution process across lineages (HAL). We also present two algorithms for searching model space and identifying an optimal model of evolution that is less likely to over- or underparameterize the data. The performance of the two new algorithms was evaluated using alignments of nucleotides with 10 000 sites simulated under complex non-SRH conditions on a 25-tipped tree. The algorithms were found to be very successful, identifying the correct HAL model with a 75% success rate (the average success rate for assigning rate matrices to the tree's 48 edges was 99.25%) and, for the correct HAL model, identifying the correct HAS model with a 98% success rate. Finally, parameter estimates obtained under the correct HAL-HAS model were found to be accurate and precise. The merits of our new algorithms were illustrated with an analysis of 42 337 second codon sites extracted from a concatenation of 106 alignments of orthologous genes encoded by the nuclear genomes of Saccharomyces cerevisiae, S. paradoxus, S. mikatae, S. kudriavzevii, S. castellii, S. kluyveri, S. bayanus, and Candida albicans. Our results show that second codon sites in the ancestral genome of these species contained 49.1% invariable sites, 39.6% variable sites belonging to one rate category (V1), and 11.3% variable sites belonging to a second rate category (V2). The ancestral nucleotide content was found to differ markedly across these three sets of sites, and the evolutionary processes operating at the variable sites were found to be non-SRH and best modeled by a combination of eight edge-specific rate matrices (four for V1 and four for V2). The number of substitutions per site at the variable sites also differed markedly, with sites belonging to V1 evolving slower than those belonging to V2 along the lineages separating the seven species of Saccharomyces. Finally, sites belonging to V1 appeared to have ceased evolving along the lineages separating S. cerevisiae, S. paradoxus, S. mikatae, S. kudriavzevii, and S. bayanus, implying that they might have become so selectively constrained that they could be considered invariable sites in these species.

Statistical tests to identify appropriate types of nucleotide sequence recoding in molecular phylogenetics.

BACKGROUND: Under a Markov model of evolution, recoding, or lumping, of the four nucleotides into fewer groups may permit analysis under simpler conditions but may unfortunately yield misleading results unless the evolutionary process of the recoded groups remains Markovian. If a Markov process is lumpable, then the evolutionary process of the recoded groups is Markovian. RESULTS: We consider stationary, reversible, and homogeneous Markov processes on two taxa and compare three tests for lumpability: one using an ad hoc test statistic, which is based on an index that is evaluated using a bootstrap approximation of its distribution; one that is based on a test proposed specifically for Markov chains; and one using a likelihood-ratio test. We show that the likelihood-ratio test is more powerful than the index test, which is more powerful than that based on the Markov chain test statistic. We also show that for stationary processes on binary trees with more than two taxa, the tests can be applied to all pairs. Finally, we show that if the process is lumpable, then estimates obtained under the recoded model agree with estimates obtained under the original model, whereas, if the process is not lumpable, then these estimates can differ substantially. We apply the new likelihood-ratio test for lumpability to two primate data sets, one with a mitochondrial origin and one with a nuclear origin. CONCLUSIONS: Recoding may result in biased phylogenetic estimates because the original evolutionary process is not lumpable. Accordingly, testing for lumpability should be done prior to phylogenetic analysis of recoded data.

Fixed, free, and fixed: the fickle phylogeny of extant Crinoidea (Echinodermata) and their Permian-Triassic origin.

Although the status of Crinoidea (sea lilies and featherstars) as sister group to all other living echinoderms is well-established, relationships among crinoids, particularly extant forms, are debated. All living species are currently placed in Articulata, which is generally accepted as the only crinoid group to survive the Permian-Triassic extinction event. Recent classifications have recognized five major extant taxa: Isocrinida, Hyocrinida, Bourgueticrinina, Comatulidina and Cyrtocrinida, plus several smaller groups with uncertain taxonomic status, e.g., Guillecrinus, Proisocrinus and Caledonicrinus. Here we infer the phylogeny of extant Crinoidea using three mitochondrial genes and two nuclear genes from 59 crinoid terminals that span the majority of extant crinoid diversity. Although there is poor support for some of the more basal nodes, and some tree topologies varied with the data used and mode of analysis, we obtain several robust results. Cyrtocrinida, Hyocrinida, Isocrinida are all recovered as clades, but two stalked crinoid groups, Bourgueticrinina and Guillecrinina, nest among the featherstars, lending support to an argument that they are paedomorphic forms. Hence, they are reduced to families within Comatulida. Proisocrinus is clearly shown to be part of Isocrinida, and Caledonicrinus may not be a bourgueticrinid. Among comatulids, tree topologies show little congruence with current taxonomy, indicating that much systematic revision is required. Relaxed molecular clock analyses with eight fossil calibration points recover Articulata with a median date to the most recent common ancestor at 231-252mya in the Middle to Upper Triassic. These analyses tend to support the hypothesis that the group is a radiation from a small clade that passed through the Permian-Triassic extinction event rather than several lineages that survived. Our tree topologies show various scenarios for the evolution of stalks and cirri in Articulata, so it is clear that further data and taxon sampling are needed to recover a more robust phylogeny of the group.

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.

Reducing model complexity of the general Markov model of evolution.

The selection of an optimal model for data analysis is an important component of model-based molecular phylogenetic studies. Owing to the large number of Markov models that can be used for data analysis, model selection is a combinatorial problem that cannot be solved by performing an exhaustive search of all possible models. Currently, model selection is based on a small subset of the available Markov models, namely those that assume the evolutionary process to be globally stationary, reversible, and homogeneous. This forces the optimal model to be time reversible even though the actual data may not satisfy these assumptions. This problem can be alleviated by including more complex models during the model selection. We present a novel heuristic that evaluates a small fraction of these complex models and identifies the optimal model.

Two stationary nonhomogeneous Markov models of nucleotide sequence evolution.

The general Markov model (GMM) of nucleotide substitution does not assume the evolutionary process to be stationary, reversible, or homogeneous. The GMM can be simplified by assuming the evolutionary process to be stationary. A stationary GMM is appropriate for analyses of phylogenetic data sets that are compositionally homogeneous; a data set is considered to be compositionally homogeneous if a statistical test does not detect significant differences in the marginal distributions of the sequences. Though the general time-reversible (GTR) model assumes stationarity, it also assumes reversibility and homogeneity. We propose two new stationary and nonhomogeneous models--one constrains the GMM to be reversible, whereas the other does not. The two models, coupled with the GTR model, comprise a set of nested models that can be used to test the assumptions of reversibility and homogeneity for stationary processes. The two models are extended to incorporate invariable sites and used to analyze a seven-taxon hominoid data set that displays compositional homogeneity. We show that within the class of stationary models, a nonhomogeneous model fits the hominoid data better than the GTR model. We note that if one considers a wider set of models that are not constrained to be stationary, then an even better fit can be obtained for the hominoid data. However, the methods for reducing model complexity from an extremely large set of nonstationary models are yet to be developed.

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.

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.

The molecular structure of the IsiA-Photosystem I supercomplex, modelled from high-resolution, crystal structures of Photosystem I and the CP43 protein.

We present the molecular structure of the IsiA-Photosystem I (PSI) supercomplex, inferred from high-resolution, crystal structures of PSI and the CP43 protein. The structure of iron-stress-induced A protein (IsiA) is similar to that of CP43, albeit with the difference that IsiA is associated with 15 chlorophylls (Chls), one more than previously assumed. The membrane-spanning helices of IsiA contain hydrophilic residues many of which bind Chl. The optimal structure of the IsiA-PSI supercomplex was inferred by systematically rearranging the IsiA monomers and PSI trimer in relation to each other. For each of the 6,969,600 structural configurations considered, we counted the number of optimal Chl-Chl connections (i.e., cases where Chl-bound Mg atoms are

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.

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.