James B. Pease

Extensive introgression in a malaria vector species complex revealed by phylogenomics

Introduction The notion that species boundaries can be porous to introgression is increasingly accepted. Yet the broader role of introgression in evolution remains contentious and poorly documented, partly because of the challenges involved in accurately identifying introgression in the very groups where it is most likely to occur. Recently diverged species often have incomplete reproductive barriers and may hybridize where they overlap. However, because of retention and stochastic sorting of ancestral polymorphisms, inference of the correct species branching order is notoriously challenging for recent speciation events, especially those closely spaced in time. Without knowledge of species relationships, it is impossible to identify instances of introgression. Rationale Since the discovery that the single mosquito taxon described in 1902 as Anopheles gambiae was actually a complex of several closely related and morphologically indistinguishable sibling species, the correct species branching order has remained controversial and unresolved. This Afrotropical complex contains the world’s most important vectors of human malaria, owing to their close association with humans, as well as minor vectors and species that do not bite humans. On the basis of ecology and behavior, one might predict phylogenetic clustering of the three highly anthropophilic vector species. However, previous phylogenetic analyses of the complex based on a limited number of markers strongly disagree about relationships between the major vectors, potentially because of historical introgression between them. To investigate the history of the species complex, we used whole-genome reference assemblies, as well as dozens of resequenced individuals from the field. Results We observed a large amount of phylogenetic discordance between trees generated from the autosomes and X chromosome. The autosomes, which make up the majority of the genome, overwhelmingly supported the grouping of the three major vectors of malaria, An. gambiae, An. coluzzii, and An. arabiensis. In stark contrast, the X chromosome strongly supported the grouping of An. arabiensis with a species that plays no role in malaria transmission, An. quadriannulatus. Although the whole-genome consensus phylogeny unequivocally agrees with the autosomal topology, we found that the topology most often located on the X chromosome follows the historical species branching order, with pervasive introgression on the autosomes producing relationships that group the three highly anthropophilic species together. With knowledge of the correct species branching order, we are further able to uncover introgression between another species pair, as well as a complex history of balancing selection, introgression, and local adaptation of a large autosomal inversion that confers aridity tolerance. Conclusion We identify the correct species branching order of the An. gambiae species complex, resolving a contentious phylogeny. Notably, lineages leading to the principal vectors of human malaria were among the first in the complex to radiate and are not most closely related to each other. Pervasive autosomal introgression between these human malaria vectors, including nonsister vector species, suggests that traits enhancing vectorial capacity can be acquired not only through de novo mutation but also through a more rapid process of interspecific genetic exchange. Time-lapse photographs of an adult anopheline mosquito emerging from its pupal case. RELATED ITEMS IN ScienceD. E. Neafsey et al., Science 347, 1258522 (2015) Introgressive hybridization is now recognized as a widespread phenomenon, but its role in evolution remains contested. Here, we use newly available reference genome assemblies to investigate phylogenetic relationships and introgression in a medically important group of Afrotropical mosquito sibling species. We have identified the correct species branching order to resolve a contentious phylogeny and show that lineages leading to the principal vectors of human malaria were among the first to split. Pervasive autosomal introgression between these malaria vectors means that only a small fraction of the genome, mainly on the X chromosome, has not crossed species boundaries. Our results suggest that traits enhancing vectorial capacity may be gained through interspecific gene flow, including between nonsister species. Mosquito adaptability across genomes Virtually everyone has first-hand experience with mosquitoes. Few recognize the subtle biological distinctions among these bloodsucking flies that render some bites mere nuisances and others the initiation of a potentially life-threatening infection. By sequencing the genomes of several mosquitoes in depth, Neafsey et al. and Fontaine et al. reveal clues that explain the mystery of why only some species of one genus of mosquitoes are capable of transmitting human malaria (see the Perspective by Clark and Messer). Science, this issue 10.1126/science.1258524 and 10.1126/science.1258522; see also p. 27 Comparison of several genomes reveals the genetic history of mosquitoes’ ability to vector malaria among humans. [Also see Perspective by Clark and Messer]

Phylogenomics Reveals Three Sources of Adaptive Variation during a Rapid Radiation

Speciation events often occur in rapid bursts of diversification, but the ecological and genetic factors that promote these radiations are still much debated. Using whole transcriptomes from all 13 species in the ecologically and reproductively diverse wild tomato clade (Solanum sect. Lycopersicon), we infer the species phylogeny and patterns of genetic diversity in this group. Despite widespread phylogenetic discordance due to the sorting of ancestral variation, we date the origin of this radiation to approximately 2.5 million years ago and find evidence for at least three sources of adaptive genetic variation that fuel diversification. First, we detect introgression both historically between early-branching lineages and recently between individual populations, at specific loci whose functions indicate likely adaptive benefits. Second, we find evidence of lineage-specific de novo evolution for many genes, including loci involved in the production of red fruit color. Finally, using a “PhyloGWAS” approach, we detect environment-specific sorting of ancestral variation among populations that come from different species but share common environmental conditions. Estimated across the whole clade, small but substantial and approximately equal fractions of the euchromatic portion of the genome are inferred to contribute to each of these three sources of adaptive genetic variation. These results indicate that multiple genetic sources can promote rapid diversification and speciation in response to new ecological opportunity, in agreement with our emerging phylogenomic understanding of the complexity of both ancient and recent species radiations.

Sex Chromosomes Evolved from Independent Ancestral Linkage Groups in Winged Insects

The evolution of a pair of chromosomes that differ in appearance between males and females (heteromorphic sex chromosomes) has occurred repeatedly across plants and animals. Recent work has shown that the male heterogametic (XY) and female heterogametic (ZW) sex chromosomes evolved independently from different pairs of homomorphic autosomes in the common ancestor of birds and mammals but also that X and Z chromosomes share many convergent molecular features. However, little is known about how often heteromorphic sex chromosomes have either evolved convergently from different autosomes or in parallel from the same pair of autosomes and how universal patterns of molecular evolution on sex chromosomes really are. Among winged insects with sequenced genomes, there are male heterogametic species in both the Diptera (e.g., Drosophila melanogaster) and the Coleoptera (Tribolium castaneum), female heterogametic species in the Lepidoptera (Bombyx mori), and haplodiploid species in the Hymenoptera (e.g., Nasonia vitripennis). By determining orthologous relationships among genes on the X and Z chromosomes of insects with sequenced genomes, we are able to show that these chromosomes are not homologous to one another but are homologous to autosomes in each of the other species. These results strongly imply that heteromorphic sex chromosomes have evolved independently from different pairs of ancestral chromosomes in each of the insect orders studied. We also find that the convergently evolved X chromosomes of Diptera and Coleoptera share genomic features with each other and with vertebrate X chromosomes, including excess gene movement from the X to the autosomes. However, other patterns of molecular evolution--such as increased codon bias, decreased gene density, and the paucity of male-biased genes on the X--differ among the insect X and Z chromosomes. Our results provide evidence for both differences and nearly universal similarities in patterns of evolution among independently derived sex chromosomes.

Powerful methods for detecting introgressed regions from population genomic data

Understanding the types and functions of genes that are able to cross species boundaries—and those that are not—is an important step in understanding the forces maintaining species as largely independent lineages across the remainder of the genome. With large next‐generation sequencing data sets we are now able to ask whether introgression has occurred across the genome, and multiple methods have been proposed to detect the signature of such events. Here, we introduce a new summary statistic that can be used to test for introgression, RNDmin, that makes use of the minimum pairwise sequence distance between two population samples relative to divergence to an outgroup. We find that our method offers a modest increase in power over other, related tests, but that all such tests have high power to detect introgressed loci when migration is recent and strong. RNDmin is robust to variation in the mutation rate, and remains reliable even when estimates of the divergence time between sister species are inaccurate. We apply RNDmin to population genomic data from the African mosquitoes Anopheles quadriannulatus and A. arabiensis, identifying three novel candidate regions for introgression. Interestingly, one of the introgressed loci is on the X chromosome, but outside of an inversion separating these two species. Our results suggest that significant, but rare, sharing of alleles is occurring between species that diverged more than 1 million years ago, and that application of these methods to additional systems are likely to reveal similar results.

More accurate phylogenies inferred from low-recombination regions in the presence of incomplete lineage sorting

When speciation events occur in rapid succession, incomplete lineage sorting (ILS) can cause disagreement among individual gene trees. The probability that ILS affects a given locus is directly related to its effective population size (Ne), which in turn is proportional to the recombination rate if there is strong selection across the genome. Based on these expectations, we hypothesized that low‐recombination regions of the genome, as well as sex chromosomes and nonrecombining chromosomes, should exhibit lower levels of ILS. We tested this hypothesis in phylogenomic datasets from primates, the Drosophila melanogaster clade, and the Drosophila simulans clade. In all three cases, regions of the genome with low or no recombination showed significantly stronger support for the putative species tree, although results from the X chromosome differed among clades. Our results suggest that recurrent selection is acting in these low‐recombination regions, such that current levels of diversity also reflect past decreases in the effective population size at these same loci. The results also demonstrate how considering the genomic context of a gene tree can assist in more accurate determination of the true species phylogeny, especially in cases where a whole‐genome phylogeny appears to be an unresolvable polytomy.

No Excess Gene Movement Is Detected off the Avian or Lepidopteran Z Chromosome

Most of our knowledge of sex-chromosome evolution comes from male heterogametic (XX/XY) taxa. With the genome sequencing of multiple female heterogametic (ZZ/ZW) taxa, we can now ask whether there are patterns of evolution common to both sex chromosome systems. In all XX/XY systems examined to date, there is an excess of testis-biased retrogenes moving from the X chromosome to the autosomes, which is hypothesized to result from either sexually antagonistic selection or escape from meiotic sex chromosome inactivation (MSCI). We examined RNA-mediated (retrotransposed) and DNA-mediated gene movement in two independently evolved ZZ/ZW systems, birds (chicken and zebra finch) and lepidopterans (silkworm). Even with sexually antagonistic selection likely operating in both taxa and MSCI having been identified in the chicken, we find no evidence for an excess of genes moving from the Z chromosome to the autosomes in either lineage. We detected no excess for either RNA- or DNA-mediated duplicates, across a range of approaches and methods. We offer some potential explanations for this difference between XX/XY and ZZ/ZW sex chromosome systems, but further work is needed to distinguish among these hypotheses. Regardless of the root causes, we have identified an additional, potentially inherent, difference between XX/XY and ZZ/ZW systems.

Quantitative Genetic Analysis Indicates Natural Selection on Leaf Phenotypes Across Wild Tomato Species (Solanum sect. Lycopersicon; Solanaceae)

Adaptive evolution requires both raw genetic material and an accessible path of high fitness from one fitness peak to another. In this study, we used an introgression line (IL) population to map quantitative trait loci (QTL) for leaf traits thought to be associated with adaptation to precipitation in wild tomatoes (Solanum sect. Lycopersicon; Solanaceae). A QTL sign test showed that several traits likely evolved under directional natural selection. Leaf traits correlated across species do not share a common genetic basis, consistent with a scenario in which selection maintains trait covariation unconstrained by pleiotropy or linkage disequilibrium. Two large effect QTL for stomatal distribution colocalized with key genes in the stomatal development pathway, suggesting promising candidates for the molecular bases of adaptation in these species. Furthermore, macroevolutionary transitions between vastly different stomatal distributions may not be constrained when such large-effect mutations are available. Finally, genetic correlations between stomatal traits measured in this study and data on carbon isotope discrimination from the same ILs support a functional hypothesis that the distribution of stomata affects the resistance to CO2 diffusion inside the leaf, a trait implicated in climatic adaptation in wild tomatoes. Along with evidence from previous comparative and experimental studies, this analysis indicates that leaf traits are an important component of climatic niche adaptation in wild tomatoes and demonstrates that some trait transitions between species could have involved few, large-effect genetic changes, allowing rapid responses to new environmental conditions.

Molecular mechanisms of postmating prezygotic reproductive isolation uncovered by transcriptome analysis

Little is known about the physiological responses and genetic mutations associated with reproductive isolation between species, especially for postmating prezygotic isolating barriers. Here, we examine changes in gene expression that accompany the expression of ‘unilateral incompatibility’ (UI)—a postmating prezygotic barrier in which fertilization is prevented by gamete rejection in the reproductive tract [in this case of pollen tubes (male gametophytes)] in one direction of a species cross, but is successful in the reciprocal crossing direction. We use whole‐transcriptome sequencing of multiple developmental stages of male and female tissues in two Solanum species that exhibit UI to: (i) identify transcript differences between UI‐competent and UI noncompetent tissues; (ii) characterize transcriptional changes specifically associated with the phenotypic expression of UI; and (iii) using these comparisons, evaluate the behaviour of a priori candidate loci for UI and identify new candidates for future manipulative work. In addition to describing transcriptome‐wide changes in gene expression that accompany this isolating barrier, we identify at least five strong candidates for involvement in postmating prezygotic incompatibility between species. These include three novel candidates and two candidates that are strongly supported by prior developmental, functional, and quantitative trait locus mapping studies. These latter genes are known molecular players in the intraspecific expression of mate choice via genetic self‐incompatibility, and our study supports prior evidence that these inter‐ and intraspecific postmating prezygotic reproductive behaviours share specific genetic and molecular mechanisms.

On the possible role of robustness in the evolution of infectious diseases

Robustness describes the capacity for a biological system to remain canalized despite perturbation. Genetic robustness affords maintenance of phenotype despite mutational input, necessarily involving the role of epistasis. Environmental robustness is phenotypic constancy in the face of environmental variation, where epistasis may be uninvolved. Here we discuss genetic and environmental robustness, from the standpoint of infectious disease evolution, and suggest that robustness may be a unifying principle for understanding how different disease agents evolve. We focus especially on viruses with RNA genomes due to their importance in the evolution of emerging diseases and as model systems to test robustness theory. We present new data on adaptive constraints for a model RNA virus challenged to evolve in response to UV radiation. We also draw attention to other infectious disease systems where robustness theory may prove useful for bridging evolutionary biology and biomedicine, especially the evolution of antibiotic resistance in bacteria, immune evasion by influenza, and malaria parasite infections.

Encoding Data Using Biological Principles: The Multisample Variant Format for Phylogenomics and Population Genomics

Rapid progress in the fields of phylogenomics and population genomics has driven increases in both the size of multi-genomic datasets and the number and complexity of genome-wide analyses. We present the Multisample Variant Format, specifically designed to store multiple sequence alignments for phylogenomics and population genomic analysis. The signature feature of MVF is a distinctive encoding of aligned sites with specific biological information content (e.g., invariant, low-coverage). This biological pattern-based encoding of sequence data allows for rapid filtering and quality control of data and speeds up computation for many analyses. Similar to other modern formats, MVF has a simple data structure and flexible header structure to accommodate project metadata, allowing to also serve as an effective data publication and sharing format. We also propose several variants of the MVF format to accommodate protein and codon alignments, quality scores, and a mix of de novo and reference-aligned data. Using the MVFtools package, MVF files can be converted from other common sequence formats. MVFtools completes tasks ranging from simple transformation and filtering operations to complex genome-wide visualizations in only a few minutes, even on large datasets. In addition to presentation of MVF and MVFtools, we also discuss the application both in MVF and other existing data formats of the broader concept of using biological principles and patterns to inform sequence data encoding.