A single evolutionary innovation drives the deep evolution of symbiotic N₂-fixation in angiosperms

Symbiotic associations occur in every habitat on earth, but we know very little about their evolutionary histories. Current models of trait evolution cannot adequately reconstruct the deep history of symbiotic innovation, because they assume homogenous evolutionary processes across millions of years. Here we use a recently developed, heterogeneous and quantitative phylogenetic framework to study the origin of the symbiosis between angiosperms and nitrogen-fixing (N2) bacterial symbionts housed in nodules. We compile the largest database of global nodulating plant species and reconstruct the symbiosis’ evolution. We identify a single, cryptic evolutionary innovation driving symbiotic N2-fixation evolution, followed by multiple gains and losses of the symbiosis, and the subsequent emergence of ‘stable fixers’ (clades extremely unlikely to lose the symbiosis). Originating over 100 MYA, this innovation suggests deep homology in symbiotic N2-fixation. Identifying cryptic innovations on the tree of life is key to understanding the evolution of complex traits, including symbiotic partnerships

Mutation Size Optimizes Speciation in an Evolutionary Model

The role of mutation rate in optimizing key features of evolutionary dynamics has recently been investigated in various computational models. Here, we address the related question of how maximum mutation size affects the formation of species in a simple computational evolutionary model. We find that the number of species is maximized for intermediate values of a mutation size parameter μ; the result is observed for evolving organisms on a randomly changing landscape as well as in a version of the model where negative feedback exists between the local population size and the fitness provided by the landscape. The same result is observed for various distributions of mutation values within the limits set by μ. When organisms with various values of μ compete against each other, those with intermediate μ values are found to survive. The surviving values of μ from these competition simulations, however, do not necessarily coincide with the values that maximize the number of species. These results suggest that various complex factors are involved in determining optimal mutation parameters for any population, and may also suggest approaches for building a computational bridge between the (micro) dynamics of mutations at the level of individual organisms and (macro) evolutionary dynamics at the species level

An Evaluation of Putative Sympatric Speciation within Limnanthes (Limnanthaceae)

Limnanthes floccosa ssp. floccosa and L. floccosa ssp. grandiflora are two of five subspecies within Limnanthes floccosa endemic to vernal pools in southern Oregon and northern California. Three seasons of monitoring natural populations have quantified that L. floccosa ssp. grandiflora is always found growing sympatrically with L. floccosa ssp. floccosa and that their flowering times overlap considerably. Despite their subspecific rank within the same species crossing experiments have confirmed that their F1 hybrids are sterile. An analysis of twelve microsatellite markers, with unique alleles in each taxon, also shows exceedingly low levels of gene flow between populations of the two subspecies. Due to the lack of previous phylogenetic resolution among L. floccosa subspecies, we used Illumina next generation sequencing to identify single nucleotide polymorphisms from genomic DNA libraries of L. floccosa ssp. floccosa and L. floccosa ssp. grandiflora. These data were used to identify single nucleotide polymorphisms in the chloroplast, mitochondrial, and nuclear genomes. From these variable loci, a total of 2772 bp was obtained using Sanger sequencing of ten individuals representing all subspecies of L. floccosa and an outgroup. The resulting phylogenetic reconstruction was fully resolved. Our results indicate that although L. floccosa ssp. floccosa and L. floccosa ssp. grandiflora are closely related, they are not sister taxa and therefore likely did not diverge as a result of a sympatric speciation event

The Repeatability of Adaptive Radiation During Long-Term Experimental Evolution of Escherichia coli in a Multiple Nutrient Environment

Adaptive radiations occur when a species diversifies into different ecological specialists due to competition for resources and trade-offs associated with the specialization. The evolutionary outcome of an instance of adaptive radiation cannot generally be predicted because chance (stochastic events) and necessity (deterministic events) contribute to the evolution of diversity. With increasing contributions of chance, the degree of parallelism among different instances of adaptive radiations and the predictability of an outcome will decrease. To assess the relative contributions of chance and necessity during adaptive radiation, we performed a selection experiment by evolving twelve independent microcosms of Escherichia coli for 1000 generations in an environment that contained two distinct resources. Specialization to either of these resources involves strong trade-offs in the ability to use the other resource. After selection, we measured three phenotypic traits: 1) fitness, 2) mean colony size, and 3) colony size diversity. We used fitness relative to the ancestor as a measure of adaptation to the selective environment; changes in colony size as a measure of the evolution of new resource specialists because colony size has been shown to correlate with resource specialization; and colony size diversity as a measure of the evolved ecological diversity. Resource competition led to the rapid evolution of phenotypic diversity within microcosms. Measurements of fitness, colony size, and colony size diversity within and among microcosms showed that the repeatability of adaptive radiation was high, despite the evolution of genetic variation within microcosms. Consistent with the observation of parallel evolution, we show that the relative contributions of chance are far smaller and less important than effects due to adaptation for the traits investigated. The two-resource environment imposed similar selection pressures in independent populations and promoted parallel phenotypic adaptive radiations in all independently evolved microcosms

Initial Mutations Direct Alternative Pathways of Protein Evolution

Whether evolution is erratic due to random historical details, or is repeatedly directed along similar paths by certain constraints, remains unclear. Epistasis (i.e. non-additive interaction between mutations that affect fitness) is a mechanism that can contribute to both scenarios. Epistasis can constrain the type and order of selected mutations, but it can also make adaptive trajectories contingent upon the first random substitution. This effect is particularly strong under sign epistasis, when the sign of the fitness effects of a mutation depends on its genetic background. In the current study, we examine how epistatic interactions between mutations determine alternative evolutionary pathways, using in vitro evolution of the antibiotic resistance enzyme TEM-1 β-lactamase. First, we describe the diversity of adaptive pathways among replicate lines during evolution for resistance to a novel antibiotic (cefotaxime). Consistent with the prediction of epistatic constraints, most lines increased resistance by acquiring three mutations in a fixed order. However, a few lines deviated from this pattern. Next, to test whether negative interactions between alternative initial substitutions drive this divergence, alleles containing initial substitutions from the deviating lines were evolved under identical conditions. Indeed, these alternative initial substitutions consistently led to lower adaptive peaks, involving more and other substitutions than those observed in the common pathway. We found that a combination of decreased enzymatic activity and lower folding cooperativity underlies negative sign epistasis in the clash between key mutations in the common and deviating lines (Gly238Ser and Arg164Ser, respectively). Our results demonstrate that epistasis contributes to contingency in protein evolution by amplifying the selective consequences of random mutations

Reciprocal Sign Epistasis between Frequently Experimentally Evolved Adaptive Mutations Causes a Rugged Fitness Landscape

The fitness landscape captures the relationship between genotype and evolutionary fitness and is a pervasive metaphor used to describe the possible evolutionary trajectories of adaptation. However, little is known about the actual shape of fitness landscapes, including whether valleys of low fitness create local fitness optima, acting as barriers to adaptive change. Here we provide evidence of a rugged molecular fitness landscape arising during an evolution experiment in an asexual population of Saccharomyces cerevisiae. We identify the mutations that arose during the evolution using whole-genome sequencing and use competitive fitness assays to describe the mutations individually responsible for adaptation. In addition, we find that a fitness valley between two adaptive mutations in the genes MTH1 and HXT6/HXT7 is caused by reciprocal sign epistasis, where the fitness cost of the double mutant prohibits the two mutations from being selected in the same genetic background. The constraint enforced by reciprocal sign epistasis causes the mutations to remain mutually exclusive during the experiment, even though adaptive mutations in these two genes occur several times in independent lineages during the experiment. Our results show that epistasis plays a key role during adaptation and that inter-genic interactions can act as barriers between adaptive solutions. These results also provide a new interpretation on the classic Dobzhansky-Muller model of reproductive isolation and display some surprising parallels with mutations in genes often associated with tumors

Genomic analysis of Acidianus hospitalis W1 a host for studying crenarchaeal virus and plasmid life cycles

The Acidianus hospitalis W1 genome consists of a minimally sized chromosome of about 2.13 Mb and a conjugative plasmid pAH1 and it is a host for the model filamentous lipothrixvirus AFV1. The chromosome carries three putative replication origins in conserved genomic regions and two large regions where non-essential genes are clustered. Within these variable regions, a few orphan orfB and other elements of the IS200/607/605 family are concentrated with a novel class of MITE-like repeat elements. There are also 26 highly diverse vapBC antitoxin–toxin gene pairs proposed to facilitate maintenance of local chromosomal regions and to minimise the impact of environmental stress. Complex and partially defective CRISPR/Cas/Cmr immune systems are present and interspersed with five vapBC gene pairs. Remnants of integrated viral genomes and plasmids are located at five intron-less tRNA genes and several non-coding RNA genes are predicted that are conserved in other Sulfolobus genomes. The putative metabolic pathways for sulphur metabolism show some significant differences from those proposed for other Acidianus and Sulfolobus species. The small and relatively stable genome of A. hospitalis W1 renders it a promising candidate for developing the first Acidianus genetic systems

Prevalence of Epistasis in the Evolution of Influenza A Surface Proteins

The surface proteins of human influenza A viruses experience positive selection to escape both human immunity and, more recently, antiviral drug treatments. In bacteria and viruses, immune-escape and drug-resistant phenotypes often appear through a combination of several mutations that have epistatic effects on pathogen fitness. However, the extent and structure of epistasis in influenza viral proteins have not been systematically investigated. Here, we develop a novel statistical method to detect positive epistasis between pairs of sites in a protein, based on the observed temporal patterns of sequence evolution. The method rests on the simple idea that a substitution at one site should rapidly follow a substitution at another site if the sites are positively epistatic. We apply this method to the surface proteins hemagglutinin and neuraminidase of influenza A virus subtypes H3N2 and H1N1. Compared to a non-epistatic null distribution, we detect substantial amounts of epistasis and determine the identities of putatively epistatic pairs of sites. In particular, using sequence data alone, our method identifies epistatic interactions between specific sites in neuraminidase that have recently been demonstrated, in vitro, to confer resistance to the drug oseltamivir; these epistatic interactions are responsible for widespread drug resistance among H1N1 viruses circulating today. This experimental validation demonstrates the predictive power of our method to identify epistatic sites of importance for viral adaptation and public health. We conclude that epistasis plays a large role in shaping the molecular evolution of influenza viruses. In particular, sites with , which would normally not be identified as positively selected, can facilitate viral adaptation through epistatic interactions with their partner sites. The knowledge of specific interactions among sites in influenza proteins may help us to predict the course of antigenic evolution and, consequently, to select more appropriate vaccines and drugs