Next‐generation phylogenetics takes root

It has been a tumultuous 5 years in phylogeography and phylogenetics during which both fields have struggled to harness the power of next‐generation sequencing (NGS) (Ekblom & Galindo 2010 ; McCormack et al. 2012a ). Fortunately, several methodological approaches appear to be taking root. In this issue of Molecular Ecology, O'Neill et al. 2013 ) employ one such method – parallel tagged sequencing (PTS) – to elucidate the phylogeography of a tiger salamander (Ambystoma tigrinum) species complex. This study demonstrates a practical application of NGS on a scale appropriate (and not overkill) for most biologists interested in phylogeography (~100 loci for ~100 individuals), and their results highlight several analytical challenges that lie ahead for researchers employing NGS techniques.     

More precisely biased: increasing the number of markers is not a silver bullet in genetic bottleneck testing

In response to our review of the use of genetic bottleneck tests in the conservation literature (Peery et al. 2012, Molecular Ecology, 21 , 3403–3418), Hoban et al. (2013, Molecular Ecology, in press) conducted population genetic simulations to show that the statistical power of genetic bottleneck tests can be increased substantially by sampling large numbers of microsatellite loci, as they suggest is now possible in the age of genomics. While we agree with Hoban and co‐workers in principle, sampling large numbers of microsatellite loci can dramatically increase the probability of committing type 1 errors (i.e. detecting a bottleneck in a stable population) when the mutation model is incorrectly assumed. Using conservative values for mutation model parameters can reduce the probability of committing type 1 errors, but doing so can result in significant losses in statistical power. Moreover, we believe that practical limitations associated with developing large numbers of high‐quality microsatellite loci continue to constrain sample sizes, a belief supported by a literature review of recent studies using next generation sequencing methods to develop microsatellite libraries. conclusion, we maintain that researchers employing genetic bottleneck tests should proceed with caution and carefully assess both statistical power and type 1 error rates associated with their study design.     

Population genomic analyses reveal possible drivers of population divergence

Recent advances in sequencing technology and efficiency enable new and improved methods to investigate how populations diverge and species evolve. Fungi have relatively small and simple genomes and can often be cultured in the laboratory. Fungal populations can thus be sequenced for a relatively low cost, which makes them ideal for population genomic analyses. In several recent population genomic studies, wild populations of fungal model organisms and human pathogens have been analysed, for example Neurospora crassa (Ellison et al. 2011 ), Saccharomyces uvarum (Almeida et al. 2014 ), Coccidioides spp. (Neafsey et al. 2010 ) and Cryptococcus gatti (Engelthaler et al. 2014 ). In this issue of Molecular Ecology, Branco et al. ( 2015 ) apply population genomic tools to understand population divergence and adaptation in a symbiotic (mycorrhizal) fungus. This study exemplifies the possibilities of diving deeper into the genomic features involved in population divergence and speciation, also for nonmodel organisms, and how molecular and analytical tools will improve our understanding of the patterns and mechanisms that underlie adaptation to habitats, population divergence and dispersal limitation of fungi.     

Is colour polymorphism advantageous to populations and species?

I am writing in response to an article by Bolton, Rollins and Griffith (2015) entitled ‘The danger within: the role of genetic, behavioural and ecological factors in population persistence of colour polymorphic species’ that was recently published as an Opinion under the NEWS AND VIEWS section in Molecular Ecology. Bolton et al. (Molecular Ecology, 2015, 24 , 2907) argue that colour polymorphism may reduce population fitness and increase extinction risk and emphasize that this is contrary to predictions put forward by Forsman et al. (Ecology, 89 , 2008, 34) and Wennersten & Forsman (Biological Reviews 87 , 2012, 756) that the existence of multiple colour morphs with co‐adapted gene complexes and associated trait values may increase the ecological and evolutionary success of polymorphic populations and species. Bolton et al. (Molecular Ecology, 2015, 24 , 2907) further state that there is no clear evidence from studies of ‘true polymorphic species’ that polymorphism promotes population persistence. In response, I (i) challenge their classifications of polymorphisms and revisit the traditional definitions recognizing the dynamic nature of polymorphisms, (ii) review empirical studies that have examined whether and how polymorphism is associated with extinction risk, (iii) discuss the roles of trait correlations between colour pattern and other phenotypic dimensions for population fitness and (iv) highlight that the causes and mechanisms that influence the composition and maintenance of polymorphisms are different from the consequences of the polymorphic condition and how it may impact on aspects of ecological success and long‐term persistence of populations and species.     

A hot topic: the genetics of adaptation to geothermal vents in Mimulus guttatus

Identifying the individual loci and mutations that underlie adaptation to extreme environments has long been a goal of evolutionary biology. However, finding the genes that underlie adaptive traits is difficult for several reasons. First, because many traits and genes evolve simultaneously as populations diverge, it is difficult to disentangle adaptation from neutral demographic processes. Second, finding the individual loci involved in any trait is challenging given the respective limitations of quantitative and population genetic methods. In this issue of Molecular Ecology, Hendrick et al. (2016) overcome these difficulties and determine the genetic basis of microgeographic adaptation between geothermal vent and nonthermal populations of Mimulus guttatus in Yellowstone National Park. The authors accomplish this by combining population and quantitative genetic techniques, a powerful, but labour‐intensive, strategy for identifying individual causative adaptive loci that few studies have used (Stinchcombe & Hoekstra 2008 ). In a previous common garden experiment (Lekberg et al. 2012), thermal M. guttatus populations were found to differ from their closely related nonthermal neighbours in various adaptive phenotypes including trichome density. Hendrick et al. (2016) combine quantitative trait loci (QTL) mapping, population genomic scans for selection and admixture mapping to identify a single genetic locus underlying differences in trichome density between thermal and nonthermal M. guttatus. The candidate gene, R2R3 MYB, is homologous to genes involved in trichome development across flowering plants. The major trichome QTL, Tr14, is also involved in trichome density differences in an independent M. guttatus population comparison (Holeski et al. 2010) making this an example of parallel genetic evolution.     

A fair fight between molecular marker types in a seascape genetics setting

From its inception, population genetics has been nearly as concerned with the genetic data type—to which analyses are brought to bear—as it is with the analysis methods themselves. The field has traversed allozymes, microsatellites, segregating sites in multilocus alignments and, currently, single nucleotide polymorphisms (SNPs) generated by high‐throughput genomic sequencing methods, primarily whole genome sequencing and reduced representation library (RRL) sequencing. As each emerging data type has gained traction, it has been compared to existing methods, based on its relative ability to discern population structural complexity at increasing levels of resolution. However, this is usually done by comparing the gold standard in one data type to the gold standard in the new data type. These gold standards frequently differ in power and in sampling density, both across a genome and throughout a spatial range. In this issue of Molecular Ecology, D’Aloia et al. apply the high‐throughput approach as fully as possible to microsatellites, nuclear loci and SNPs genotyped through an RRL method; this is coupled with a spatially dense sampling scheme. Completing a battery of population genetics analyses across data types (including a series of down‐sampled data sets), the authors find that SNP data are slightly more sensitive to fine‐scale genetic structure, and the results are more resilient to down‐sampling than microsatellites and nonrepetitive nuclear loci. However, their results are far from an unqualified victory for RRL SNP data over all previous data types: the authors note that modest additions to the microsatellites and nuclear loci data sets may provide the necessary analytical power to delineate the fine‐scale genetic structuring identified by SNPs. As always, as the field begins to fully embrace the newest thing, good science reminds us that traditional data types are far from useless, especially when combined with a well‐designed sampling scheme.     

Metagenome skimming for phylogenetic community ecology: a new era in biodiversity research

It is now well recognized that considering species evolutionary history is crucial for understanding the processes driving community assembly (Cavender‐Bares et al. 2009 ). Considerable efforts have been made to integrate phylogenetics and community ecology into a single theoretical framework. Yet, assessing phylogenetic structure at the community scale remains a great challenge, in particular for poorly known organisms. While DNA metabarcoding is increasingly used for assessing taxonomic composition of complex communities from environmental samples, biases and limitations of this technique can preclude the retrieval of information on phylogenetic community structure. In this issue of Molecular Ecology, Andújar et al. (2015) demonstrate that shotgun sequencing of bulk samples of soil beetles and subsequent reconstruction of mitochondrial genomes can provide a solid phylogenetic framework to estimate species diversity and gain insights into the mechanisms underlying the spatial turnover of soil mesofaunal assemblages. This work highlights the enormous potential of ‘metagenome skimming’ not only for improving the current standards of DNA‐based biodiversity assessment but also for opening up the application of phylogenetic community ecology to hyperdiverse and poorly known biota, which was heretofore inconceivable.     

Next‐generation sequencing sheds light on intricate regulation of insect gut microbiota

Next‐generation sequencing (NGS) technologies are getting cheaper and easier and hence becoming readily accessible for many researchers in biological disciplines including ecology. In this issue of Molecular Ecology, Sudakaran et al. (2012) show how the NGS revolution contributes to our better and more comprehensive understanding of ecological interactions between gut symbiotic microbiota and the host organism. Using the European red firebug Pyrrhocoris apterus as a model system, they demonstrated that the gut microbiota consists of a small number of major bacterial phylotypes plus other minor bacterial associates. The major bacteria are localized in a specific anoxic section of the midgut and quantitatively account for most of the gut microbiota irrespective of host's geographic populations. The specific gut microbiota is established through early nymphal development of the host insect. Interestingly, the host feeding on different food, namely linden seeds, sunflower seeds or wasp larvae, scarcely affected the symbiont composition, suggesting homoeostatic control over the major symbiotic microbiota in the anoxic section of the midgut. Some of the minor components of the gut microbiota, which conventional PCR/cloning/sequencing approaches would have failed to detect, were convincingly shown to be food‐derived. These findings rest on the robust basis of high‐throughput sequencing data, and some of them could not be practically obtained by conventional molecular techniques, highlighting the significant impact of NGS approaches on ecological aspects of host–symbiont interactions in a nonmodel organism.     

Standing and flowing: the complex origins of adaptive variation

A population faced with a new selection pressure can only adapt if appropriate genetic variation is available. This genetic variation might come from new mutations or from gene exchange with other populations or species, or it might already segregate in the population as standing genetic variation (which might itself have arisen from either mutation or gene flow). Understanding the relative importance of these sources of adaptive variation is a fundamental issue in evolutionary genetics (Orr & Betancourt 2001 ; Barrett & Schluter 2008 ; Gladyshev et al. 2008 ) and has practical implications for conservation, plant and animal breeding, biological control and infectious disease prevention (e.g. Robertson 1960 ; Soulé & Wilcox 1980 ; Prentis et al. 2008 ; Pennings 2012 ). In this issue of Molecular Ecology, Roesti et al. ( 2014 ) make an important contribution to this longstanding debate.     

Will reduced host connectivity curb the spread of a devastating epidemic?

The white‐nose syndrome (WNS), caused by the fungal pathogen Pseudogymnoascus destructans, is threatening the cave‐dwelling bat fauna of North America by killing individuals by the thousands in hibernacula each winter since its appearance in New York State less than ten years ago. Epidemiological models predict that WNS will reach the western coast of the USA by 2035, potentially eliminating most populations of susceptible bat species in its path (Frick et al. 2015; O'Regan et al. 2015). These models were built and validated using distributional data from the early years of the epidemic, which spread throughout eastern North America following a route driven by cave density and winter severity (Maher et al. 2012). In this issue of Molecular Ecology, Wilder et al. (2015) refine these findings by showing that connectivity among host populations, as assessed by population genetic markers, is crucial in determining the spread of the pathogen. Because host connectivity is much reduced in the hitherto disease free western half of North America, Wilder et al. make the reassuring prediction that the disease will spread more slowly west of the Great Plains.     

Ecological genomics in full colour

Colour patterns in animals have long offered an opportunity to observe adaptive traits in natural populations. Colour plays myriad roles in interactions within and among species, from reproductive signalling to predator avoidance, leading to multiple targets of natural and sexual selection and opportunities for diversification. Understanding the genetic and developmental underpinnings of variation in colour promises a fuller understanding of these evolutionary processes, but the path to unravelling these connections can be arduous. The advent of genomic techniques suitable for nonmodel organisms is now beginning to light the way. Two new studies in this issue of Molecular Ecology use genomic sequencing of laboratory crosses to map colour traits in cichlid fishes, a remarkably diverse group in which coloration has played a major role in diversification. They illustrate how genomic approaches, specifically RAD sequencing, can rapidly identify both simple and more complex genetic variation underlying ecologically important traits. In the first, Henning et al. ( 2014 ) detect a single locus that appears to control in a Mendelian fashion the presence of horizontal stripes, a trait that has evolved in numerous cichlid lineages. In the second, Albertson et al. ( 2014 ) identify several genes and epistatic interactions affecting multiple colour traits, as well as a novel metric describing integration across colour traits. Albertson et al. ( 2014 ) go further, by quantifying differential expression of parental alleles at a candidate locus and by relating differentiation among natural populations at mapped loci to trait divergence. Herein lies the promise of ecological genomics – efficiently integrating genetic mapping of phenotypes with population genomic data to both identify functional genes and unravel their evolutionary history. These studies offer guidance on how genomic techniques can be tailored to a research question or study system, and they also add to the growing body of empirical examples addressing basic questions about how ecologically important traits evolve in natural populations.     

Ancient hyaenas highlight the old problem of estimating evolutionary rates

Phylogenetic analyses of ancient DNA data can provide a timeline for evolutionary change even in the absence of fossils. The power to infer the evolutionary rate is, however, highly dependent on the number and age of samples, the information content of the sequence data and the demographic history of the sampled population. In this issue of Molecular Ecology, Sheng et al. ( 2014 ) analysed mitochondrial DNA sequences isolated from a combination of ancient and present‐day hyaenas, including three Pleistocene samples from China. Using an evolutionary rate inferred from the ages of the ancient sequences, they recalibrated the timing of hyaena diversification and suggest a much more recent evolutionary history than was believed previously. Their results highlight the importance of accurately estimating the evolutionary rate when inferring timescales of geographical and evolutionary diversification.     

Mapping the genomic architecture of ecological speciation in the wild: does linkage disequilibrium hold the key?

The hunt for the genes underlying ecological speciation has now closed in on a number of candidates, but making the link from genotype to phenotype continues to pose a significant challenge. This is partly because genetic studies in many systems remain impeded by long generation times or an inability to perform controlled crosses. Now, in this issue of Molecular Ecology, Malek et al. (2012) demonstrate the utility of a novel admixture mapping approach that can be used to identify genomic regions contributing to adaptive trait divergence between natural populations. Remarkably, they validate their approach by mapping traits associated with mate choice in a wild limnetic and benthic threespine stickleback (Gasterosteus aculeatus) species pair, finding several loci associated with male nuptial coloration and shape. While this study benefited from tried‐and‐true microsatellites in a well‐characterized species with a detailed genetic map (and genome sequence), the field is quickly moving towards the use of next‐generation sequencing, especially for nonmodel systems. The ability to characterize molecular polymorphisms for any system suggests that molecular ecologists working on virtually any species may benefit from applying Malek et al.'s approach, if naturally admixed populations are available.     

Coalescing molecular evolution and DNA barcoding

The DNA barcoding concept (Woese et al. 1990 ; Hebert et al. 2003 ) has considerably boosted taxonomy research by facilitating the identification of specimens and discovery of new species. Used alone or in combination with DNA metabarcoding on environmental samples (Taberlet et al. 2012 ), the approach is becoming a standard for basic and applied research in ecology, evolution and conservation across taxa, communities and ecosystems (Scheffers et al. 2012 ; Kress et al. 2015 ). However, DNA barcoding suffers from several shortcomings that still remain overlooked, especially when it comes to species delineation (Collins & Cruickshank 2012 ). In this issue of Molecular Ecology, Barley & Thomson ( 2016 ) demonstrate that the choice of models of sequence evolution has substantial impacts on inferred genetic distances, with a propensity of the widely used Kimura 2‐parameter model to lead to underestimated species richness. While DNA barcoding has been and will continue to be a powerful tool for specimen identification and preliminary taxonomic sorting, this work calls for a systematic assessment of substitution models fit on barcoding data used for species delineation and reopens the debate on the limitation of this approach.     

Army ant invasions reveal phylogeographic processes across the Isthmus of Panama

Female army ants cannot fly, making them very poor dispersers across water barriers. This dependence on terrestrial corridors motivated the investigation by Winston et al. ( 2017 ), published in this issue of Molecular Ecology, into the role of Panamanian isthmus formation in the diversification of Eciton army ants. Complete closure of this isthmus occurred around three million years ago (3 Ma), but it has also been hypothesized that earlier, temporary land connections facilitated additional colonization events between South and Central America over the past 13 million years or more. The phylogenomic and population genomic analyses by Winston et al. ( 2017 ) uncovered multiple incursions of Eciton lineages into Central America between 4 and 7 Ma. Their study contributes to a growing body of evidence arguing that transitory land bridges predating 3 Ma supported substantial intercontinental biotic exchange.     

Genome scans for the contemporary response to selection in quantitative traits

Genome scans have been an important approach for discovering historical signatures of selection in both model and nonmodel species. An exciting new experimental design for genome scans is to measure the change in allele frequency before and after contemporary selection within a generation, from a single population. The most widely‐used methods, however, have two major limitations: they are based on testing one locus at a time, and they only have power to uncover loci that have evolved under relatively strong selection. On the other hand, complex quantitative traits are common in nature and are caused by several loci of small effect. Selection on a quantitative trait at the phenotypic level is predicted to be accompanied by subtle allele frequency changes in many loci that covary (a polygenic soft sweep), rather than a large, single‐effect allele (a selective sweep). In this issue of Molecular Ecology, Bourret et al. (2014) measure the contemporary response to natural selection across the genome in multiple cohorts of Atlantic salmon during their first year at sea. They introduce a multilocus framework based on groups of markers that covary in their genotypic distribution. While the traditional, single‐locus approach did not find evidence for repeated patterns of selection, the multivariate approach found that a group of covarying SNPs was selected for in different cohorts at one site. Their multilocus framework has potential to be a more fruitful approach for uncovering the genomic basis of adaptation in quantitative traits, although caution should be applied as the framework has yet to be validated with simulated data.     

Don't throw the baby out with the bathwater: identifying and mapping paralogs in salmonids

Many eukaryotic genomes contain a large fraction of gene duplicates (or paralogs) as a result of ancient or recent whole‐genome duplications (Ohno 1970 ; Jaillon et al. 2004 ; Kellis et al. 2004 ). Identifying paralogs with NGS data is a pervasive problem in both ancient polyploids and neopolyploids. Likewise, paralogs are often treated as a nuisance that has to be detected and removed (Everett et al. 2012 ). In this issue of Molecular Ecology Resources, Waples et al. ( 2015 ) show that exclusion might not be necessary and how we may miss out on important genomic information in doing so. They present a novel statistical approach to detect paralogs based on the segregation of RAD loci in haploid offspring and test their method by constructing linkage maps with and without these duplicated loci in chum salmon, Oncorhynchus keta (Fig.  1 ). Their linkage map including the resolved paralogs shows that these are mostly located in the distal regions of several linkage groups. Particularly intriguing is their finding that these homoeologous regions appear impoverished in transposable elements (TE). Given the role that TE play in genome remodelling, it is noteworthy that these elements are of low abundance in regions showing residual tetrasomic inheritance. This raises the question whether re‐diploidization is constrained in these regions and whether they might have a role to play in salmonid speciation. This study provides an original approach to identifying duplicated loci in species with a pedigree, as well as providing a dense linkage map for chum salmon, and interesting insights into the retention of gene duplicates in an ancient polyploid.     

Novel aspects in the life cycle and biotrophic interactions in Pezizomycetes (Ascomycota,Fungi)

The ascomycete class Pezizomycetes (single order Pezizales) is known for its cup‐shaped fruit bodies and the evolution of edible truffles and morels, but little is known about the ontogeny and ecology of this large and ecologically diverse fungal group. In this issue of Molecular Ecology, Healy et al. ( 2013 ) make a great leap forward by describing and identifying asexual, anamorphic structures that produce mitotic spores in many ectomycorrhiza‐forming truffle and nontruffle species on soil surfaces worldwide (Fig.  1 ). Although such anamorphic forms have been reported sporadically from certain ectomycorrhizal and saprotrophic Pezizomycetes (e.g. Warcup 1990 ), Healy et al. ( 2013 ) demonstrate that these terricolous asexual forms are both taxonomically and geographically more widespread and, in fact, much more common than previously understood. We anticipate that deeper insight into other substrates, provided by molecular analyses of materials such as dead wood and seeds, is likely to reveal numerous anamorphs of saprotrophic and pathogenic Pezizomycetes as well (see Marek et al. 2009 ).     

The discovery of Antarctic RNA viruses: a new game changer

Antarctic ecosystems are dominated by micro‐organisms, and viruses play particularly important roles in the food webs. Since the first report in 2009 (López‐Bueno et al. 2009 ), ‘omic’‐based studies have greatly enlightened our understanding of Antarctic aquatic microbial diversity and ecosystem function (Wilkins et al. 2013 ; Cavicchioli 2015 ). This has included the discovery of many new eukaryotic viruses (López‐Bueno et al. 2009 ), virophage predators of algal viruses (Yau et al. 2011 ), bacteria with resistance to phage (Lauro et al. 2011 ) and mechanisms of haloarchaeal evasion, defence and adaptation to viruses (Tschitschko et al. 2015 ). In this issue of Molecular Ecology, López‐Bueno et al. ( 2015 ) report the first discovery of RNA viruses from an Antarctic aquatic environment. High sequence coverage enabled genome variation to be assessed for four positive‐sense single‐stranded RNA viruses from the order Picornavirales. By examining the populations present in the water column and in the lake's catchment area, populations of ‘quasispecies’ were able to be linked to local environmental factors. In view of the importance of viruses in Antarctic ecosystems but lack of data describing them, this study represents a significant advance in the field.