Site Frequency Spectrum Research Articles

Effect of the Unfolded Protein Response and Oxidative Stress on Mutagenesis of CSF3R a Model of Evolution of Severe Congenital Neutropenia to Myelodysplastic Syndrome/Acute Myeloid Leukemia

Background. Severe congenital neutropenia (SCN) is an inherited bone marrow failure syndrome that can transform to myelodysplastic syndrome/acute myeloid leukemia (MDS/AML). The most common recurrent mutation that causes SCN involve neutrophil elastase (ELANE). Mutations in ELANE result in the unfolded protein response stress. The treatment of choice for SCN is the chronic administration of high-dose granulocyte-colony stimulating factor (G-CSF), which elevates the neutrophil count, helps resolve pre-existing infections, diminishes the number of new infections, and significantly improves the survival and quality of life. G-CSF treatment also leads to enhanced oxidative stress. Long-term survival with G-CSF is frequently associated with development of MDS/AML. Of note, approximately 70% of SCN patients with MDS/AML acquire nonsense mutations in the region of CSF3R that encodes the cytoplasmic domain. These somatic CSF3R mutations are characterized by a truncation variant found in the cytoplasmic domain of the CSF3R and are associated with a hyper-proliferative/impaired differentiative phenotype that might contribute to the MDS/AML transformation. We hypothesize that the terminal exon of CSF3R constitutes a hotspot vulnerable to mutations resulting from excessive oxidative stress or endoplasmic reticulum (ER) stress. Results. Murine factor-dependent Ba/F3 cells were used to measure the effect of induced oxidative or ER stress on the mutation rate involving the hypothesized hotspot of the exogenous human CSF3R, the corresponding region in the endogenous Csf3r, and Runx1 (a transcription factor involved in leukemogenesis). Ba/F3 cells transduced with the cDNA for the hypothesized hotspot of CSF3R (partial C-terminal) fused in-frame with m-NeonGreen, a yellow-green fluorescent protein, were subjected to stress-inducing treatment for 30 days (~51 doubling times). The amplicon-based targeted deep-sequencing data for days 15 and 30 samples showed increased mutagenesis of the coding nucleotide sequences for CSF3R, Csf3r, and Runx1. There was no correlation between the stress-inducing chemical treatments and overall level of mutagenesis in Ba/F3 cells. Interestingly, the GC-rich partial CSF3R region was less mutated as compared to the mNeonGreen region, having much lower GC content. However, analysis of our data, including the site-frequency spectra, indicated effects that may be due to clonal selection, specifically at the Csf3r gene. Conclusion. Our data suggested that oxidative or ER stress induction did not promote genomic instability affecting the 3' exonic end of CSF3R, the endogenous Csf3R, and the endogenous Runx1 in Ba/F3 cells that could account for these targets being mutational hotspots. We conclude that other mechanisms, such as stochastic events, result in mutations of CSF3R drive the evolution of SCN to MDS/AML. Disclosures No relevant conflicts of interest to declare.

Genome-wide heritability analysis of severe malaria resistance reveals evidence of polygenic inheritance.

Estimating single nucleotide polymorphism (SNP)-heritability (h2g) of severe malaria resistance and its distribution across the genome might shed new light in to the underlying biology. We investigated h2g of severe malaria resistance from a genome-wide association study (GWAS) dataset (sample size = 11 657). We estimated the h2g and partitioned in to chromosomes, allele frequencies and annotations using the genetic relationship-matrix restricted maximum likelihood approach. We further examined non-cell type-specific and cell type-specific enrichments from GWAS-summary statistics. The h2g of severe malaria resistance was estimated at 0.21 (se = 0.05, P = 2.7 × 10-5), 0.20 (se = 0.05, P = 7.5 × 10-5) and 0.17 (se = 0.05, P = 7.2 × 10-4) in Gambian, Kenyan and Malawi populations, respectively. A comparable range of h2g [0.21 (se = 0.02, P < 1 × 10-5)] was estimated from GWAS-summary statistics meta-analysed across the three populations. Partitioning analysis from raw genotype data showed significant enrichment of h2g in genic SNPs while summary statistics analysis suggests evidences of enrichment in multiple categories. Supporting the polygenic inheritance, the h2g of severe malaria resistance is distributed across the chromosomes and allelic frequency spectrum. However, the h2g is disproportionately concentrated on three chromosomes (chr 5, 11 and 20), suggesting cost-effectiveness of targeting these chromosomes in future malaria genomic sequencing studies. We report for the first time that the heritability of malaria resistance is largely ascribed by common SNPs and the causal variants are overrepresented in protein coding regions of the genome. Further studies with larger sample sizes are needed to better understand the underpinning genetics of severe malaria resistance.

Comparison of the Full Distribution of Fitness Effects of New Amino Acid Mutations Across Great Apes.

The distribution of fitness effects (DFE) is central to many questions in evolutionary biology. However, little is known about the differences in DFE between closely related species. We use >9000 coding genes orthologous one-to-one across great apes, gibbons, and macaques to assess the stability of the DFE across great apes. We use the unfolded site frequency spectrum of polymorphic mutations (n = 8 haploid chromosomes per population) to estimate the DFE. We find that the shape of the deleterious DFE is strikingly similar across great apes. We confirm that effective population size (Ne ) is a strong predictor of the strength of negative selection, consistent with the nearly neutral theory. However, we also find that the strength of negative selection varies more than expected given the differences in Ne between species. Across species, mean fitness effects of new deleterious mutations covaries with Ne , consistent with positive epistasis among deleterious mutations. We find that the strength of negative selection for the smallest populations, bonobos and western chimpanzees, is higher than expected given their Ne This may result from a more efficient purging of strongly deleterious recessive variants in these populations. Forward simulations confirm that these findings are not artifacts of the way we are inferring Ne and DFE parameters. All findings are replicated using only GC-conservative mutations, thereby confirming that GC-biased gene conversion is not affecting our conclusions.

A Single-Couple Human Origin is Possible

The problem of inferring history from genetic data is complex and underdetermined; there are many possible scenarios that would explain the same data. It can be made more tractable by making reasonable simplifications to the model, but it is continually important to remember what has been demonstrated and what is merely a parsimonious working assumption. In this paper we have chosen to model the demographic ancestry of humanity using the simplest of assumptions, with a homogeneous population whose size can vary over time. All other assumptions such as the mutation rates were standard, and no natural selection was in operation. Using a previously published backwards simulation method and some newly developed and faster algorithms, we run our single-couple origin model of humanity and compare the results to allele frequency spectra and linkage disequilibrium statistics from current genetic data. We show that a single-couple origin of humanity as recent as 500kya is consistent with data. With only minor modifications of our parsimonious model assumptions, we suggest that a single-couple origin 100kya, or more recently, is possible.

Parsing the synonymous mutations in the maize genome: isoaccepting mutations are more advantageous in regions with codon co-occurrence bias

BackgroundSynonymous mutations do not change amino acids but do sometimes change the tRNAs (anticodons) that decode a particular codon. An isoaccepting codon is a synonymous codon that shares the same tRNA. If a mutated codon could base pair with the same anticodon as the original, the mutation is termed an isoaccepting mutation. An interesting but less-studied type of codon bias is codon co-occurrence bias. There is a trend to cluster the isoaccepting codons in the genome. The proposed advantage of codon co-occurrence bias is that the tRNA released from the ribosome E site could be quickly recharged and subsequently decode the following isoaccepting codons. This advantage would enhance translation efficiency. In plant species, whether there are signals of positive selection on isoaccepting mutations in the codon co-occurred regions has not been studied.ResultsWe termed polymorphic mutations in coding regions using publicly available RNA-seq data in maize (Zea mays). Next, we classified all synonymous mutations into three categories according to the context, i.e., the relationship between the focal codon and the previous codon, as follows: isoaccepting, nonisoaccepting and nonsynonymous. We observed higher fractions of isoaccepting mutations in the isoaccepting context. If we looked at the minor allele frequency (MAF) spectrum, the isoaccepting mutations have a higher MAF in the isoaccepting context than that in other regions, and accordingly, the nonisoaccepting mutations have a higher MAF in the nonisoaccepting context.ConclusionOur results indicate that in regions with codon co-occurrence bias, natural selection maintains this pattern by suppressing the nonisoaccepting mutations. However, if the consecutive codons are nonisoaccepting, mutations tend to switch these codons to become isoaccepting. Our study demonstrates that the codon co-occurrence bias in the maize genome is selectively maintained by natural selection and that the advantage of this trend could potentially be the rapid recharging and reuse of tRNAs to increase translation efficiency.

Joint imputation of whole-genome sequence variants and large chromosomal deletions in cattle

Genotype imputation, often focused on SNP and small insertions and deletions (indels; size ≤50 bp), is a crucial step for association mapping and estimation of genomic breeding values. Here, we present strategies to impute genotypes for large chromosomal deletions (size >50 bp), along with SNP and indels in cattle. The pipelines include a strategy for extending the whole-genome sequence reference panel for large deletions, a 2-step genotype refinement approach using Beagle4 and SHAPEIT2 software, and finally, joint imputation of SNP, indels, and large deletions to the existing SNP array-typed population using Minimac3 software. Using these pipelines we achieved an imputation accuracy of the squared Pearson correlation (r2) > 0.6 at minor allele frequencies as low as 0.7% for SNP and indels, and 0.2% for large deletions. This highlights the potential of our approach to build a haplotype reference panel and impute different classes of sequence variants across a wide allele frequency spectrum with high accuracy.

Evaluating Population Genetic Structure and Demographic History of Quercus spinosa (Fagaceae) Based on Specific Length Amplified Fragment Sequencing.

Effectively identifying the genetic structure and related factors of a species can facilitate understanding the evolutionary history of the species. Phylogeographic patterns and genetic data are essential in investigating the species historical processes and diversification that response to environmental, climatic and geological influences. In this study, Specific Length Amplified Fragment Sequencing (SLAF-seq) data and ecological niche models (ENMs) are combined to identify the genetic structure and demographic modeling of Quercus spinosa, and evaluate the impacts of historical range shifts, climatic variation, and landscape factors on this species. The population topology and genetic divergence of the Cenozoic were inferred by a site frequency spectrum based composite-likelihood approach which is a novel strategy for maximizing the utility of linked SLAF markers. The overall genetic structure using model-based and model-free clustering methods was consistently identified as two geographically distinct genetic clusters. A deep divergence between two natural lineages (i.e., a western Himalaya-Hengduan Mountains lineage and an eastern Qin-ling Mountains lineage) was observed. The demographic modeling and Niche reconstruction indicated that the two groups were diverged in the late Miocene and then presented as two distinct genetic lineages. With the Quaternary glacial climate fluctuation, two groups had continuous asymmetrical secondary contact and gene exchange in the Sichuan Basin during the last glacial maximum. Besides, a significant relationship between genetic distance and geography in all individuals was identified by the Mantel test. Overall, this study 1) contributes to a better understanding of the role played by Quaternary climatic fluctuation in the present-day distributions of Q. spinosa; 2) provides a comprehensive view of the genome-wide variation of sclerophyllous forests in ecological adaptive evolution; 3) indicates that dispersal limitation and ecological divergence contribute to the genome-wide differentiation of Q. spinosa, which supports a hypothesis that complex geography and climatic changes strongly influence the evolutionary origin and history of the species.

Tree lengths for general $\Lambda $-coalescents and the asymptotic site frequency spectrum around the Bolthausen–Sznitman coalescent

We study tree lengths in $\Lambda $-coalescents without a dust component from a sample of $n$ individuals. For the total length of all branches and the total length of all external branches, we present laws of large numbers in full generality. The other results treat regularly varying coalescents with exponent 1, which cover the Bolthausen–Sznitman coalescent. The theorems contain laws of large numbers for the total length of all internal branches and of internal branches of order $a$ (i.e., branches carrying $a$ individuals out of the sample). These results immediately transform to sampling formulas in the infinite sites model. In particular, we obtain the asymptotic site frequency spectrum of the Bolthausen–Sznitman coalescent. The proofs rely on a new technique to obtain laws of large numbers for certain functionals of decreasing Markov chains.

Extensive disruption of protein interactions by genetic variants across the allele frequency spectrum in human populations

Each human genome carries tens of thousands of coding variants. The extent to which this variation is functional and the mechanisms by which they exert their influence remains largely unexplored. To address this gap, we leverage the ExAC database of 60,706 human exomes to investigate experimentally the impact of 2009 missense single nucleotide variants (SNVs) across 2185 protein-protein interactions, generating interaction profiles for 4797 SNV-interaction pairs, of which 421 SNVs segregate at > 1% allele frequency in human populations. We find that interaction-disruptive SNVs are prevalent at both rare and common allele frequencies. Furthermore, these results suggest that 10.5% of missense variants carried per individual are disruptive, a higher proportion than previously reported; this indicates that each individual’s genetic makeup may be significantly more complex than expected. Finally, we demonstrate that candidate disease-associated mutations can be identified through shared interaction perturbations between variants of interest and known disease mutations.

Synaptic and Gene Regulatory Mechanisms in Schizophrenia, Autism, and 22q11.2 Copy Number Variant–Mediated Risk for Neuropsychiatric Disorders

Background22q11.2 copy number variants are among the most highly penetrant genetic risk variants for developmental neuropsychiatric disorders such as schizophrenia (SCZ) and autism spectrum disorder (ASD). However, the specific mechanisms through which they confer risk remain unclear. MethodsUsing a functional genomics approach, we integrated transcriptomic data from the developing human brain, genome-wide association findings for SCZ and ASD, protein interaction data, and gene expression signatures from SCZ and ASD postmortem cortex to 1) organize genes into the developmental cellular and molecular systems within which they operate, 2) identify neurodevelopmental processes associated with polygenic risk for SCZ and ASD across the allelic frequency spectrum, and 3) elucidate pathways and individual genes through which 22q11.2 copy number variants may confer risk for each disorder. ResultsPolygenic risk for SCZ and ASD converged on partially overlapping neurodevelopmental modules involved in synaptic function and transcriptional regulation, with ASD risk variants additionally enriched for modules involved in neuronal differentiation during fetal development. The 22q11.2 locus formed a large protein network during development that disproportionately affected SCZ-associated and ASD-associated neurodevelopmental modules, including loading highly onto synaptic and gene regulatory pathways. SEPT5, PI4KA, and SNAP29 genes are candidate drivers of 22q11.2 synaptic pathology relevant to SCZ and ASD, and DGCR8 and HIRA are candidate drivers of disease-relevant alterations in gene regulation. ConclusionsThis approach offers a powerful framework to identify neurodevelopmental processes affected by diverse risk variants for SCZ and ASD and elucidate mechanisms through which highly penetrant, multigene copy number variants contribute to disease risk.

Genotyping Method and Frequency of ADRB3-rs4994 Single Nucleotide Polymorphism Genotypes in Hanoi 3-5 Years Old Chidren

Genotyping Method and Frequency of ADRB3-rs4994 Single Nucleotide Polymorphism Genotypes in Hanoi 3-5 Years Old Chidren

Historical demography of the Caribbean spiny lobster Panulirus argus (Latreille, 1804) (Decapoda: Achelata: Palinuridae) in the Florida Keys, USA inferred using single nucleotide polymorphisms (SNPs)

ABSTRACT The Caribbean spiny lobster Panulirus argus (Latreille, 1804) is an important species in shallow-water coral reefs and target of the most lucrative fishery in the Caribbean Sea. We explored historical demography in P. argus inferred using single nucleotide polymorphisms (SNPs). We expected an increase in population size of P. argus from Florida, USA starting ~18,000–24,000 years ago, after the Last Glacial Maximum, when ice sheets started to retreat and sub-tropical/tropical shallow coastal waters warmed up. A total of 10 lobsters were collected from shallow reefs in the Florida Keys, Florida, USA. One microgram of gDNA extracted from each specimen was used for RAD library construction using established protocols. A panel of 1643 SNPs obtained after interrogation of RAD-tags was used to calculate a site frequency spectrum (SFS). The observed SFS for the Florida population of P. argus exhibited a non-normal distribution peaking at singleton SNPs. The expected SFS in a total of six different candidate demographic models with dissimilar population size changes through time (i.e., standard neutral, exponential growth, bottleneck, bottleneck + growth, two epochs, and three epochs) were numerically computed in the software ∂a∂i and a model selection approach was implemented to test which expected model(s) best fitted the empirical SFS. The model selection approach indicated that the bottleneck + growth model most closely matched the observed SFS; P. argus experienced a population decline at about 1.9 (0.75–5.7) mya, to then recover and growth exponentially until present time. In disagreement with expectations, population expansion started much earlier than ~18,000–24,000 years ago. Fisheries and conservation studies are expected to profit from the evaluation of genomic and population variability in this species using demographic models, as shown here. Studies exploring population connectivity and locality-specific demographic history of P. argus are underway.

Population Genomic Signatures of Genetic Structure and Environmental Selection in the Catadromous Roughskin Sculpin Trachidermus fasciatus.

Understanding the patterns of genetic diversity and adaptation across species’ range is crucial to assess its long-term persistence and determine appropriate conservation measures. The impacts of human activities on the genetic diversity and genetic adaptation to heterogeneous environments remain poorly understood in the marine realm. The roughskin sculpin (Trachidermus fasciatus) is a small catadromous fish, and has been listed as a second-class state protected aquatic animal since 1988 in China. To elucidate the underlying mechanism of population genetic structuring and genetic adaptations to local environments, RAD tags were sequenced for 202 individuals in nine populations across the range of T. fasciatus in China. The pairwise FST values over 9,271 filtered SNPs were significant except that between Dongying and Weifang. All the genetic clustering analysis revealed significant population structure with high support for eight distinct genetic clusters. Both the minor allele frequency spectra and Ne estimations suggested extremely small Ne in some populations (e.g., Qinhuangdao, Rongcheng, Wendeng, and Qingdao), which might result from recent population bottleneck. The strong genetic structure can be partly attributed to genetic drift and habitat fragmentation, likely due to the anthropogenic activities. Annotations of candidate adaptive loci suggested that genes involved in metabolism, development, and osmoregulation were critical for adaptation to spatially heterogenous environment of local populations. In the context of anthropogenic activities and environmental change, results of the present population genomic work provided important contributions to the understanding of genetic differentiation and adaptation to changing environments.

Genomic reconstruction of 100 000-year grassland history in a forested country: population dynamics of specialist forbs.

Grassland ecosystems worldwide have been extensively converted to other land uses and are globally imperiled. Because many grasslands have been maintained by human activities, understanding their origin and history is fundamentally important to better contemporary management. However, existing methods to reconstruct past vegetation can produce contrasting views on grassland history. Here, we inferred demographic histories of 40 populations of four grassland forb species throughout Japan using high-resolution genome sequences and model-flexible demographic simulation based on the site frequency spectrum. Although two species showed a slight decline in population size between 100 000–10 000 years ago, our results suggest that population sizes of studied species have been maintained within the range of 0.5–2.0 times the most recent estimates for at least 100 000 years across Japan. Our results suggest that greater than 90% declines in Japanese grasslands and subsequent losses of grassland species in the last 100 years are geologically and biologically important and will have substantial consequences for Japanese biota and culture. People have had critical roles in maintaining disturbance-dependent grassland ecosystems and biota in this warm and wet forested country. In these contexts, disturbances associated with forest harvesting and traditional extensive farming have the potential to maintain grassland ecosystems and can provide important opportunities to reconcile resource production and conservation of grassland biodiversity.

Pleistocene climate changes, and not agricultural spread, accounts for range expansion and admixture in the dominant grassland species Lolium perenne L.

AbstractAimGrasslands have been pivotal in the development of herbivore breeding since the Neolithic and still represent the most widespread agricultural land use across Europe. However, it remains unclear whether the current large‐scale genetic variation of plant species found in natural grasslands of Europe is the result of human activities or natural processes.LocationEurope.TaxonLolium perenne L. (perennial ryegrass).MethodsWe reconstructed the phylogeographic history of L. perenne, a dominant grassland species, using 481 natural populations, including 11 populations of closely related taxa. We combined Genotyping‐by‐Sequencing (GBS) and pool‐Sequencing (pool‐Seq) to obtain high‐quality allele frequency calls of ~500 k SNP loci. We performed genetic structure analyses and demographic reconstructions based on the site frequency spectrum (SFS). We additionally used the same genotyping protocol to assess the genomic diversity of a set of 32 cultivars representative of the L. perenne cultivars widely used for forage purposes.ResultsExpansion across Europe took place during the Würm glaciation (12–110 kya), a cooling period that decreased the dominance of trees in favour of grasses. Splits and admixtures in L. perenne fit historical climate changes in the Mediterranean basin. The development of agriculture in Europe (7–3.5 kya), that caused an increase in the abundance of grasslands, did not have an effect on the demographic patterns of L. perenne. We found that most modern cultivars are closely related to natural diversity from north‐western Europe. Thus, modern cultivars do not represent the wide genetic variation found in natural populations.Main conclusionsDemographic events in L. perenne can be explained by the changing climatic conditions during the Pleistocene. Natural populations maintain a wide genomic variability at continental scale that has been minimally exploited by recent breeding activities. This variability constitutes valuable standing genetic variation for future adaptation of grasslands to climate change, safeguarding the agricultural services they provide.

Model-based demographic inference of introgression history in European whitefish species pairs'.

Parallel phenotypic differentiation is generally attributed to parallel adaptive divergence as an evolutionary response to similar environmental contrasts. Such parallelism may actually originate from several evolutionary scenarios ranging from repeated parallel divergence caused by divergent selection to a unique divergence event followed by gene flow. Reconstructing the evolutionary history underlying parallel phenotypic differentiation is thus fundamental to understand the relative contribution of demography and selection on genomic divergence during speciation. In this study, we investigate the divergence history of replicate European whitefish (Coregonus lavaretus), limnetic and benthic species pairs from two lakes in Norway and two lakes in Switzerland. Demographic models accounting for semi-permeability and linked selection were fitted to the unfolded joint allele frequency spectrum built from genome-wide SNPs and compared to each other in each species pair. We found strong support for a model of asymmetrical post-glacial secondary contact between glacial lineages in all four lakes. Moreover, our results suggest that heterogeneous genomic differentiation has been shaped by the joint action of linked selection accelerating lineage sorting during allopatry, and heterogeneous migration eroding divergence at different rates along the genome following secondary contact. Our analyses reveal how the interplay between demography, selection and historical contingency has influenced the levels of diversity observed in previous whitefish phylogeographic studies. This study thus provides new insights into the historical demographic and selective processes that shaped the divergence associated with ecological speciation in European whitefish.

Evolutionary genomics of gypsy moth populations sampled along a latitudinal gradient.

The European gypsy moth (Lymantria dispar L.) was first introduced to Massachusetts in 1869 and within 150years has spread throughout eastern North America. This large-scale invasion across a heterogeneous landscape allows examination of the genetic signatures of adaptation potentially associated with rapid geographical spread. We tested the hypothesis that spatially divergent natural selection has driven observed changes in three developmental traits that were measured in a common garden for 165 adult moths sampled from six populations across a latitudinal gradient covering the entirety of the range. We generated genotype data for 91,468 single nucleotide polymorphisms based on double digest restriction-site associated DNA sequencing and used these data to discover genome-wide associations for each trait, as well as to test for signatures of selection on the discovered architectures. Genetic structure across the introduced range of gypsy moth was low in magnitude (FST =0.069), with signatures of bottlenecks and spatial expansion apparent in the rare portion of the allele frequency spectrum. Results from applications of Bayesian sparse linear mixed models were consistent with the presumed polygenic architectures of each trait. Further analyses indicated spatially divergent natural selection acting on larval development time and pupal mass, with the linkage disequilibrium component of this test acting as the main driver of observed patterns. The populations most important for these signals were two range-edge populations established less than 30 generations ago. We discuss the importance of rapid polygenic adaptation to the ability of non-native species to invade novel environments.

A Log-Ratio Biplot Approach for Exploring Genetic Relatedness Based on Identity by State.

The detection of cryptic relatedness in large population-based cohorts is of great importance in genome research. The usual approach for detecting closely related individuals is to plot allele sharing statistics, based on identity-by-state or identity-by-descent, in a two-dimensional scatterplot. This approach ignores that allele sharing data across individuals has in reality a higher dimensionality, and neither regards the compositional nature of the underlying counts of shared genotypes. In this paper we develop biplot methodology based on log-ratio principal component analysis that overcomes these restrictions. This leads to entirely new graphics that are essentially useful for exploring relatedness in genetic databases from homogeneous populations. The proposed method can be applied in an iterative manner, acting as a looking glass for more remote relationships that are harder to classify. Datasets from the 1,000 Genomes Project and the Genomes For Life-GCAT Project are used to illustrate the proposed method. The discriminatory power of the log-ratio biplot approach is compared with the classical plots in a simulation study. In a non-inbred homogeneous population the classification rate of the log-ratio principal component approach outperforms the classical graphics across the whole allele frequency spectrum, using only identity by state. In these circumstances, simulations show that with 35,000 independent bi-allelic variants, log-ratio principal component analysis, combined with discriminant analysis, can correctly classify relationships up to and including the fourth degree.

Genotype-free estimation of allele frequencies reduces bias and improves demographic inference from RADSeq data.

Restriction-site associated DNA sequencing (RADSeq) facilitates rapid generation of thousands of genetic markers at relatively low cost; however, several sources of error specific to RADSeq methods often lead to biased estimates of allele frequencies and thereby to erroneous population genetic inference. Estimating the distribution of sample allele frequencies without calling genotypes was shown to improve population inference from whole genome sequencing data, but the ability of this approach to account for RADSeq-specific biases remains unexplored. Here we assess in how far genotype-free methods of allele frequency estimation affect demographic inference from empirical RADSeq data. Using the well-studied pied flycatcher (Ficedula hypoleuca) as a study system, we compare allele frequency estimation and demographic inference from whole genome sequencing data with that from RADSeq data matched for samples using both genotype-based and genotype free methods. The demographic history of pied flycatchers as inferred from RADSeq data was highly congruent with that inferred from whole genome resequencing (WGS) data when allele frequencies were estimated directly from the read data. In contrast, when allele frequencies were derived from called genotypes, RADSeq-based estimates of most model parameters fell outside the 95% confidence interval of estimates derived from WGS data. Notably, more stringent filtering of the genotype calls tended to increase the discrepancy between parameter estimates from WGS and RADSeq data, respectively. The results from this study demonstrate the ability of genotype-free methods to improve allele frequency spectrum- (AFS-) based demographic inference from empirical RADSeq data and highlight the need to account for uncertainty in NGS data regardless of sequencing method.

The Impact of Natural Selection on Short Insertion and Deletion Variation in the Great Tit Genome.

Insertions and deletions (INDELs) remain understudied, despite being the most common form of genetic variation after single nucleotide polymorphisms. This stems partly from the challenge of correctly identifying the ancestral state of an INDEL and thus identifying it as an insertion or a deletion. Erroneously assigned ancestral states can skew the site frequency spectrum, leading to artificial signals of selection. Consequently, the selective pressures acting on INDELs are, at present, poorly resolved. To tackle this issue, we have recently published a maximum likelihood approach to estimate the mutation rate and the distribution of fitness effects for INDELs. Our approach estimates and controls for the rate of ancestral state misidentification, overcoming issues plaguing previous INDEL studies. Here, we apply the method to INDEL polymorphism data from ten high coverage (∼44×) European great tit (Parus major) genomes. We demonstrate that coding INDELs are under strong purifying selection with a small proportion making it into the population (∼4%). However, among fixed coding INDELs, 71% of insertions and 86% of deletions are fixed by positive selection. In noncoding regions, we estimate ∼80% of insertions and ∼52% of deletions are effectively neutral, the remainder show signatures of purifying selection. Additionally, we see evidence of linked selection reducing INDEL diversity below background levels, both in proximity to exons and in areas of low recombination.