Adaptive evolution of the enigmatic Takakia now facing climate change in Tibet.

The most extreme environments are the most vulnerable to transformation under a rapidly changing climate. These ecosystems harbor some of the most specialized species, which will likely suffer the highest extinction rates. We document the steepest temperature increase (2010-2021) on record at altitudes of above 4,000 m, triggering a decline of the relictual and highly adapted moss Takakia lepidozioides. Its de-novo-sequenced genome with 27,467 protein-coding genes includes distinct adaptations to abiotic stresses and comprises the largest number of fast-evolving genes under positive selection. The uplift of the study site in the last 65 million years has resulted in life-threatening UV-B radiation and drastically reduced temperatures, and we detected several of the molecular adaptations of Takakia to these environmental changes. Surprisingly, specific morphological features likely occurred earlier than 165 mya in much warmer environments. Following nearly 400 million years of evolution and resilience, this species is now facing extinction.

Modeling the mosaic structure of bacterial genomes to infer their evolutionary history.

The chronology and phylogeny of bacterial evolution are difficult to reconstruct due to a scarce fossil record. The analysis of bacterial genomes remains challenging because of large sequence divergence, the plasticity of bacterial genomes due to frequent gene loss, horizontal gene transfer, and differences in selective pressure from one locus to another. Therefore, taking advantage of the rich and rapidly accumulating genomic data requires accurate modeling of genome evolution. An important technical consideration is that loci with high effective mutation rates may diverge beyond the detection limit of the alignment algorithms used, biasing the genome-wide divergence estimates toward smaller divergences. In this article, we propose a novel method to gain insight into bacterial evolution based on statistical properties of genome comparisons. We find that the length distribution of sequence matches is shaped by the effective mutation rates of different loci, by the horizontal transfers, and by the aligner sensitivity. Based on these inputs, we build a model and show that it accounts for the empirically observed distributions, taking the Enterobacteriaceae family as an example. Our method allows to distinguish segments of vertical and horizontal origins and to estimate the time divergence and exchange rate between any pair of taxa from genome-wide alignments. Based on the estimated time divergences, we construct a time-calibrated phylogenetic tree to demonstrate the accuracy of the method.

Genomes, fossils, and the concurrent rise of modern birds and flowering plants in the Late Cretaceous.

The phylogeny and divergence timing of the Neoavian radiation remain controversial despite recent progress. We analyzed the genomes of 124 species across all Neoavian orders, using data from 25,460 loci spanning four DNA classes, including 5,756 coding sequences, 12,449 conserved nonexonic elements, 4,871 introns, and 2,384 intergenic segments. We conducted a comprehensive sensitivity analysis to account for the heterogeneity across different DNA classes, leading to an optimal tree of Neoaves with high resolution. This phylogeny features a novel Neoavian dichotomy comprising two monophyletic clades: a previously recognized Telluraves (land birds) and a newly circumscribed Aquaterraves (waterbirds and relatives). Molecular dating analyses with 20 fossil calibrations indicate that the diversification of modern birds began in the Late Cretaceous and underwent a constant and steady radiation across the KPg boundary, concurrent with the rise of angiosperms as well as other major Cenozoic animal groups including placental and multituberculate mammals. The KPg catastrophe had a limited impact on avian evolution compared to the Paleocene-Eocene Thermal Maximum, which triggered a rapid diversification of seabirds. Our findings suggest that the evolution of modern birds followed a slow process of gradualism rather than a rapid process of punctuated equilibrium, with limited interruption by the KPg catastrophe. This study places bird evolution into a new context within vertebrates, with ramifications for the evolution of the Earth's biota.

Dating Ammonia-Oxidizing Bacteria with Abundant Eukaryotic Fossils.

Evolution of a complete nitrogen (N) cycle relies on the onset of ammonia oxidation, which aerobically converts ammonia to nitrogen oxides. However, accurate estimation of the antiquity of ammonia-oxidizing bacteria (AOB) remains challenging because AOB-specific fossils are absent and bacterial fossils amenable to calibrate molecular clocks are rare. Leveraging the ancient endosymbiosis of mitochondria and plastid, as well as using state-of-the-art Bayesian sequential dating approach, we obtained a timeline of AOB evolution calibrated largely by eukaryotic fossils. We show that the first AOB evolved in marine Gammaproteobacteria (Gamma-AOB) and emerged between 2.1 and 1.9 billion years ago (Ga), thus postdating the Great Oxidation Event (GOE; 2.4 to 2.32 Ga). To reconcile the sedimentary N isotopic signatures of ammonia oxidation occurring near the GOE, we propose that ammonia oxidation likely occurred at the common ancestor of Gamma-AOB and Gammaproteobacterial methanotrophs, or the actinobacterial/verrucomicrobial methanotrophs which are known to have ammonia oxidation activities. It is also likely that nitrite was transported from the terrestrial habitats where ammonia oxidation by archaea took place. Further, we show that the Gamma-AOB predated the anaerobic ammonia-oxidizing (anammox) bacteria, implying that the emergence of anammox was constrained by the availability of dedicated ammonia oxidizers which produce nitrite to fuel anammox. Our work supports a new hypothesis that N redox cycle involving nitrogen oxides evolved rather late in the ocean.

Loss of endogenous estrogen alters mitochondrial metabolism and muscle clock-related protein Rbm20 in female mdx mice.

Female carriers of a Duchenne muscular dystrophy (DMD) gene mutation manifest exercise intolerance and metabolic anomalies that may be exacerbated following menopause due to the loss of estrogen, a known regulator of skeletal muscle function and metabolism. Here, we studied the impact of estrogen depletion (via ovariectomy) on exercise tolerance and muscle mitochondrial metabolism in female mdx mice and the potential of estrogen replacement therapy (using estradiol) to protect against functional and metabolic perturbations. We also investigated the effect of estrogen depletion, and replacement, on the skeletal muscle proteome through an untargeted proteomic approach with TMT-labelling. Our study confirms that loss of estrogen in female mdx mice reduces exercise capacity, tricarboxylic acid cycle intermediates, and citrate synthase activity but that these deficits are offset through estrogen replacement therapy. Furthermore, ovariectomy downregulated protein expression of RNA-binding motif factor 20 (Rbm20), a critical regulator of sarcomeric and muscle homeostasis gene splicing, which impacted pathways involving ribosomal and mitochondrial translation. Estrogen replacement modulated Rbm20 protein expression and promoted metabolic processes and the upregulation of proteins involved in mitochondrial dynamics and metabolism. Our data suggest that estrogen mitigates dystrophinopathic features in female mdx mice and that estrogen replacement may be a potential therapy for post-menopausal DMD carriers.

Clockor2: Inferring global and local strict molecular clocks using root-to-tip regression.

Molecular sequence data from rapidly evolving organisms are often sampled at different points in time. Sampling times can then be used for molecular clock calibration. The root-to-tip (RTT) regression is an essential tool to assess the degree to which the data behave in a clock-like fashion. Here, we introduce Clockor2, a client-side web application for conducting RTT regression. Clockor2 allows users to quickly fit local and global molecular clocks, thus handling the increasing complexity of genomic datasets that sample beyond the assumption of homogeneous host populations. Clockor2 is efficient, handling trees of up to the order of 104 tips, with significant speed increases compared to other RTT regression applications. Although clockor2 is written as a web application, all data processing happens on the client-side, meaning that data never leaves the user's computer. Clockor2 is freely available at https : //clockor2.github.io/.

LSM12 and ME31B/DDX6 Define Distinct Modes of Posttranscriptional Regulation by ATAXIN-2 Protein Complex in Drosophila Circadian Pacemaker Neurons.

ATAXIN-2 (ATX2) has been implicated in human neurodegenerative diseases, yet it remains elusive how ATX2 assembles specific protein complexes to execute its physiological roles. Here we employ the posttranscriptional co-activator function of Drosophila ATX2 to demonstrate that LSM12 and ME31B/DDX6 are two ATX2-associating factors crucial for sustaining circadian rhythms. LSM12 acts as a molecular adaptor for the recruitment of TWENTY-FOUR (TYF) to ATX2. The ATX2-LSM12-TYF complex thereby stimulates TYF-dependent translation of the rate-limiting clock gene period (per) to maintain 24 hr periodicity in circadian behaviors. In contrast, ATX2 contributes to NOT1-mediated gene silencing and associates with NOT1 in a ME31B/DDX6-dependent manner. The ME31B/DDX6-NOT1 complex does not affect PER translation but supports high-amplitude behavioral rhythms along with ATX2, indicating a PER-independent clock function of ATX2. Taken together, these data suggest that the ATX2 complex may switch distinct modes of posttranscriptional regulation through its associating factors to control circadian clocks and ATX2-related physiology.

Two light sensors decode moonlight versus sunlight to adjust a plastic circadian/circalunidian clock to moon phase.

Many species synchronize their physiology and behavior to specific hours. It is commonly assumed that sunlight acts as the main entrainment signal for ∼24-h clocks. However, the moon provides similarly regular time information. Consistently, a growing number of studies have reported correlations between diel behavior and lunidian cycles. Yet, mechanistic insight into the possible influences of the moon on ∼24-h timers remains scarce. We have explored the marine bristleworm Platynereis dumerilii to investigate the role of moonlight in the timing of daily behavior. We uncover that moonlight, besides its role in monthly timing, also schedules the exact hour of nocturnal swarming onset to the nights' darkest times. Our work reveals that extended moonlight impacts on a plastic clock that exhibits <24 h (moonlit) or >24 h (no moon) periodicity. Abundance, light sensitivity, and genetic requirement indicate that the Platynereis light receptor molecule r-Opsin1 serves as a receptor that senses moonrise, whereas the cryptochrome protein L-Cry is required to discriminate the proper valence of nocturnal light as either moonlight or sunlight. Comparative experiments in Drosophila suggest that cryptochrome's principle requirement for light valence interpretation is conserved. Its exact biochemical properties differ, however, between species with dissimilar timing ecology. Our work advances the molecular understanding of lunar impact on fundamental rhythmic processes, including those of marine mass spawners endangered by anthropogenic change.

Fossil-calibrated molecular clock data enable reconstruction of steps leading to differentiated multicellularity and anisogamy in the Volvocine algae.

BACKGROUND: Throughout its nearly four-billion-year history, life has undergone evolutionary transitions in which simpler subunits have become integrated to form a more complex whole. Many of these transitions opened the door to innovations that resulted in increased biodiversity and/or organismal efficiency. The evolution of multicellularity from unicellular forms represents one such transition, one that paved the way for cellular differentiation, including differentiation of male and female gametes. A useful model for studying the evolution of multicellularity and cellular differentiation is the volvocine algae, a clade of freshwater green algae whose members range from unicellular to colonial, from undifferentiated to completely differentiated, and whose gamete types can be isogamous, anisogamous, or oogamous. To better understand how multicellularity, differentiation, and gametes evolved in this group, we used comparative genomics and fossil data to establish a geologically calibrated roadmap of when these innovations occurred. RESULTS: Our ancestral-state reconstructions, show that multicellularity arose independently twice in the volvocine algae. Our chronograms indicate multicellularity evolved during the Carboniferous-Triassic periods in Goniaceae + Volvocaceae, and possibly as early as the Cretaceous in Tetrabaenaceae. Using divergence time estimates we inferred when, and in what order, specific developmental changes occurred that led to differentiated multicellularity and oogamy. We find that in the volvocine algae the temporal sequence of developmental changes leading to differentiated multicellularity is much as proposed by David Kirk, and that multicellularity is correlated with the acquisition of anisogamy and oogamy. Lastly, morphological, molecular, and divergence time data suggest the possibility of cryptic species in Tetrabaenaceae. CONCLUSIONS: Large molecular datasets and robust phylogenetic methods are bringing the evolutionary history of the volvocine algae more sharply into focus. Mounting evidence suggests that extant species in this group are the result of two independent origins of multicellularity and multiple independent origins of cell differentiation. Also, the origin of the Tetrabaenaceae-Goniaceae-Volvocaceae clade may be much older than previously thought. Finally, the possibility of cryptic species in the Tetrabaenaceae provides an exciting opportunity to study the recent divergence of lineages adapted to live in very different thermal environments.

Timing without coding: How do long non-coding RNAs regulate circadian rhythms?

Long non-coding RNAs (lncRNAs) are a new class of regulatory RNAs that play important roles in disease development and a variety of biological processes. Recent studies have underscored the importance of lncRNAs in the circadian clock system and demonstrated that lncRNAs regulate core clock genes and the core clock machinery in mammals. In this review, we provide an overview of our current understanding of how lncRNAs regulate the circadian clock without coding a protein. We also offer additional insights into the challenges in understanding the functions of lncRNAs and other unresolved questions in the field. We do not cover other regulatory ncRNAs even though they also play important roles; readers are highly encouraged to refer to other excellent reviews on this topic.

More than the clock: distinct regulation of muscle function and metabolism by PER2 and RORα.

Circadian rhythms, governed by the dominant central clock, in addition to various peripheral clocks, regulate almost all biological processes, including sleep-wake cycles, hormone secretion and metabolism. In certain contexts, the regulation and function of the peripheral oscillations can be decoupled from the central clock. However, the specific mechanisms underlying muscle-intrinsic clock-dependent modulation of muscle function and metabolism remain unclear. We investigated the outcome of perturbations of the primary and secondary feedback loops of the molecular clock in skeletal muscle by specific gene ablation of Period circadian regulator 2 (Per2) and RAR-related orphan receptor alpha (Rorα), respectively. In both models, a dampening of core clock gene oscillation was observed, while the phase was preserved. Moreover, both loops seem to be involved in the homeostasis of amine groups. Highly divergent outcomes were seen for overall muscle gene expression, primarily affecting circadian rhythmicity in the PER2 knockouts and non-oscillating genes in the RORα knockouts, leading to distinct outcomes in terms of metabolome and phenotype. These results highlight the entanglement of the molecular clock and muscle plasticity and allude to specific functions of different clock components, i.e. the primary and secondary feedback loops, in this context. The reciprocal interaction between muscle contractility and circadian clocks might therefore be instrumental to determining a finely tuned adaptation of muscle tissue to perturbations in health and disease. KEY POINTS: Specific perturbations of the primary and secondary feedback loop of the molecular clock result in specific outcomes on muscle metabolism and function. Ablation of Per2 (primary loop) or Rorα (secondary loop) blunts the amplitude of core clock genes, in absence of a shift in phase. Perturbation of the primary feedback loop by deletion of PER2 primarily affects muscle gene oscillation. Knockout of RORα and the ensuing modulation of the secondary loop results in the aberrant expression of a large number of non-clock genes and proteins. The deletion of PER2 and RORα affects muscle metabolism and contractile function in a circadian manner, highlighting the central role of the molecular clock in modulating muscle plasticity.

Detecting Episodic Evolution through Bayesian Inference of Molecular Clock Models.

Molecular evolutionary rate variation is a key aspect of the evolution of many organisms that can be modeled using molecular clock models. For example, fixed local clocks revealed the role of episodic evolution in the emergence of SARS-CoV-2 variants of concern. Like all statistical models, however, the reliability of such inferences is contingent on an assessment of statistical evidence. We present a novel Bayesian phylogenetic approach for detecting episodic evolution. It consists of computing Bayes factors, as the ratio of posterior and prior odds of evolutionary rate increases, effectively quantifying support for the effect size. We conducted an extensive simulation study to illustrate the power of this method and benchmarked it to formal model comparison of a range of molecular clock models using (log) marginal likelihood estimation, and to inference under a random local clock model. Quantifying support for the effect size has higher sensitivity than formal model testing and is straight-forward to compute, because it only needs samples from the posterior and prior distribution. However, formal model testing has the advantage of accommodating a wide range molecular clock models. We also assessed the ability of an automated approach, known as the random local clock, where branches under episodic evolution may be detected without their a priori definition. In an empirical analysis of a data set of SARS-CoV-2 genomes, we find "very strong" evidence for episodic evolution. Our results provide guidelines and practical methods for Bayesian detection of episodic evolution, as well as avenues for further research into this phenomenon.

Epimutations Define a Fast-Ticking Molecular Clock in Plants.

Stochastic gains and losses of DNA methylation at CG dinucleotides are a frequent occurrence in plants. These spontaneous 'epimutations' occur at a rate that is 100 000 times higher than the genetic mutation rate, are effectively neutral at the genome-wide scale, and are stably inherited across mitotic and meiotic cell divisions. Mathematical models have been extraordinarily successful at describing how epimutations accumulate in plant genomes over time, making this process one of the most predictable epigenetic phenomena to date. Here, we propose that their high rate and effective neutrality make epimutations a powerful new molecular clock for timing evolutionary events of the recent past and for age dating of long-lived perennials such as trees.

Diversity and biogeography of South American mud turtles elucidated by multilocus DNA sequencing (Testudines: Kinosternidae).

Kinosternon is the most speciose genus of extant turtles, with 22 currently recognized species, distributed across large parts of the Americas. Most species have small distributions, but K. leucostomum and K. scorpioides range from Mexico to South America. Previous studies have found discordance between mitochondrial and nuclear phylogenies in some kinosternid groups, with the current taxonomy following the nuclear-based results. Herein, based on extended molecular, geographic, and taxonomic sampling, we explore the phylogeographic structure and taxonomic limits for K. leucostomum and the K. scorpioides group and present a fossil-calibrated nuclear time tree for Kinosternon. Our results reveal contrasting differentiation patterns for the K. scorpioides group and K. leucostomum, despite overlapping distributions. Kinosternon leucostomum shows only shallow geographic divergence, whereas the K. scorpioides group is polyphyletic with up to 10 distinct taxa, some of them undescribed. We support the elevation of K. s. albogulare and K. s. cruentatum to species level. Given the deep divergence within the genus Kinosternon, we propose the recognition of three subgenera, Kinosternon, Cryptochelys and Thyrosternum, and the abandonment of the group-based classification, at least for the K. leucostomum and K. scorpioides groups. Our results show an initial split in Kinosternon that gave rise to two main radiations, one Nearctic and one mainly Neotropical. Most speciation events in Kinosternon occurred during the Quaternary and we hypothesize that they were mediated by both climatic and geological events. Additionally, our data imply that at least three South American colonizations occurred, two in the K. leucostomum group, and one in the K. scorpioides group. Additionally, we hypothesize that discordance between mitochondrial and nuclear phylogenetic signal is due to mitochondrial capture from an extinct kinosternine lineage.

Phylogeny and biogeography of harmochirine jumping spiders (Araneae: Salticidae).

We use ultraconserved elements (UCE) and Sanger data to study the phylogeny, age, and biogeographical history of harmochirine jumping spiders, a group that includes the species-rich genus Habronattus, whose remarkable courtship has made it the focus of studies of behaviour, sexual selection, and diversification. We recovered 1947 UCE loci from 43 harmochirine taxa and 4 outgroups, yielding a core dataset of 193 UCEs with at least 50 % occupancy. Concatenated likelihood and ASTRAL analyses confirmed the separation of harmochirines into two major clades, here designated the infratribes Harmochirita and Pellenita. Most are African or Eurasian with the notable exception of a clade of pellenites containing Habronattus and Pellenattus of the Americas and Havaika and Hivanua of the Pacific Islands. Biogeographical analysis using the DEC model favours a dispersal of the clade's ancestor from Eurasia to the Americas, from which Havaika's ancestor dispersed to Hawaii and Hivanua's ancestor to the Marquesas Islands. Divergence time analysis on 32 loci with 85 % occupancy, calibrated by fossils and island age, dates the dispersal to the Americas at approximately 4 to 6 million years ago. The explosive radiation of Habronattus perhaps began only about 4 mya. The phylogeny clarifies both the evolution of sexual traits (e.g., the terminal apophyses was enlarged in Pellenes and not subsequently lost) and the taxonomy. Habronattus is confirmed as monophyletic. Pellenattus is raised to the status of genus, and 13 species moved into it as new combinations. Bianor stepposus Logunov, 1991 is transferred to Sibianor, and Pellenes bulawayoensis Wesolowska, 1999 is transferred to Neaetha. A molecular clock rate estimate for spider UCEs is presented and its utility to inform prior distributions is discussed.

Evaluating the Accuracy of Methods for Detecting Correlated Rates of Molecular and Morphological Evolution.

Determining the link between genomic and phenotypic change is a fundamental goal in evolutionary biology. Insights into this link can be gained by using a phylogenetic approach to test for correlations between rates of molecular and morphological evolution. However, there has been persistent uncertainty about the relationship between these rates, partly because conflicting results have been obtained using various methods that have not been examined in detail. We carried out a simulation study to evaluate the performance of 5 statistical methods for detecting correlated rates of evolution. Our simulations explored the evolution of molecular sequences and morphological characters under a range of conditions. Of the methods tested, Bayesian relaxed-clock estimation of branch rates was able to detect correlated rates of evolution correctly in the largest number of cases. This was followed by correlations of root-to-tip distances, Bayesian model selection, independent sister-pairs contrasts, and likelihood-based model selection. As expected, the power to detect correlated rates increased with the amount of data, both in terms of tree size and number of morphological characters. Likewise, greater among-lineage rate variation in the data led to improved performance of all 5 methods, particularly for Bayesian relaxed-clock analysis when the rate model was mismatched. We then applied these methods to a data set from flowering plants and did not find evidence of a correlation in evolutionary rates between genomic data and morphological characters. The results of our study have practical implications for phylogenetic analyses of combined molecular and morphological data sets, and highlight the conditions under which the links between genomic and phenotypic rates of evolution can be evaluated quantitatively.

Drosophila clock cells use multiple mechanisms to transmit time-of-day signals in the brain.

Regulation of circadian behavior and physiology by the Drosophila brain clock requires communication from central clock neurons to downstream output regions, but the mechanism by which clock cells regulate downstream targets is not known. We show here that the pars intercerebralis (PI), previously identified as a target of the morning cells in the clock network, also receives input from evening cells. We determined that morning and evening clock neurons have time-of-day-dependent connectivity to the PI, which is regulated by specific peptides as well as by fast neurotransmitters. Interestingly, PI cells that secrete the peptide DH44, and control rest:activity rhythms, are inhibited by clock inputs while insulin-producing cells (IPCs) are activated, indicating that the same clock cells can use different mechanisms to drive cycling in output neurons. Inputs of morning cells to IPCs are relevant for the circadian rhythm of feeding, reinforcing the role of the PI as a circadian relay that controls multiple behavioral outputs. Our findings provide mechanisms by which clock neurons signal to nonclock cells to drive rhythms of behavior.

Angiosperm-wide analysis of fruit and ovary evolution aided by a new nuclear phylogeny supports association of the same ovary type with both dry and fleshy fruits.

Fruit functions in seed protection and dispersal and belongs to many dry and fleshy types, yet their evolutionary pattern remains unclear in part due to uncertainties in the phylogenetic relationships among several orders and families. Thus we used nuclear genes of 502 angiosperm species representing 231 families to reconstruct a well supported phylogeny, with resolved relationships for orders and families with previously uncertain placements. Using this phylogeny as a framework, molecular dating supports a Triassic origin of the crown angiosperms, followed by the emergence of most orders in the Jurassic and Cretaceous and their rise to ecological dominance during the Cretaceous Terrestrial Revolution. The robust phylogeny allowed an examination of the evolutionary pattern of fruit and ovary types, revealing a trend of parallel carpel fusions during early diversifications in eudicots, monocots, and magnoliids. Moreover, taxa in the same order or family with the same ovary type can develop either dry or fleshy fruits with strong correlations between specific types of dry and fleshy fruits; such associations of ovary, dry and fleshy fruits define several ovary-fruit "modules" each found in multiple families. One of the frequent modules has an ovary containing multiple ovules, capsules and berries, and another with an ovary having one or two ovules, achenes (or other single-seeded dry fruits) and drupes. This new perspective of relationships among fruit types highlights the closeness of specific dry and fleshy fruit types, such as capsule and berry, that develop from the same ovary type and belong to the same module relative to dry and fleshy fruits of other modules (such as achenes and drupes). Further analyses of gene families containing known genes for ovary and fruit development identified phylogenetic nodes with multiple gene duplications, supporting a possible role of whole-genome duplications, in combination with climate changes and animal behaviors, in angiosperm fruit and ovary diversification.

Purifying Selection Determines the Short-Term Time Dependency of Evolutionary Rates in SARS-CoV-2 and pH1N1 Influenza.

High-throughput sequencing enables rapid genome sequencing during infectious disease outbreaks and provides an opportunity to quantify the evolutionary dynamics of pathogens in near real-time. One difficulty of undertaking evolutionary analyses over short timescales is the dependency of the inferred evolutionary parameters on the timespan of observation. Crucially, there are an increasing number of molecular clock analyses using external evolutionary rate priors to infer evolutionary parameters. However, it is not clear which rate prior is appropriate for a given time window of observation due to the time-dependent nature of evolutionary rate estimates. Here, we characterize the molecular evolutionary dynamics of SARS-CoV-2 and 2009 pandemic H1N1 (pH1N1) influenza during the first 12 months of their respective pandemics. We use Bayesian phylogenetic methods to estimate the dates of emergence, evolutionary rates, and growth rates of SARS-CoV-2 and pH1N1 over time and investigate how varying sampling window and data set sizes affect the accuracy of parameter estimation. We further use a generalized McDonald-Kreitman test to estimate the number of segregating nonneutral sites over time. We find that the inferred evolutionary parameters for both pandemics are time dependent, and that the inferred rates of SARS-CoV-2 and pH1N1 decline by ∼50% and ∼100%, respectively, over the course of 1 year. After at least 4 months since the start of sequence sampling, inferred growth rates and emergence dates remain relatively stable and can be inferred reliably using a logistic growth coalescent model. We show that the time dependency of the mean substitution rate is due to elevated substitution rates at terminal branches which are 2-4 times higher than those of internal branches for both viruses. The elevated rate at terminal branches is strongly correlated with an increasing number of segregating nonneutral sites, demonstrating the role of purifying selection in generating the time dependency of evolutionary parameters during pandemics.

The Emergence of SARS-CoV-2 Variants of Concern Is Driven by Acceleration of the Substitution Rate.

The ongoing SARS-CoV-2 pandemic has seen an unprecedented amount of rapidly generated genome data. These data have revealed the emergence of lineages with mutations associated to transmissibility and antigenicity, known as variants of concern (VOCs). A striking aspect of VOCs is that many of them involve an unusually large number of defining mutations. Current phylogenetic estimates of the substitution rate of SARS-CoV-2 suggest that its genome accrues around two mutations per month. However, VOCs can have 15 or more defining mutations and it is hypothesized that they emerged over the course of a few months, implying that they must have evolved faster for a period of time. We analyzed genome sequence data from the GISAID database to assess whether the emergence of VOCs can be attributed to changes in the substitution rate of the virus and whether this pattern can be detected at a phylogenetic level using genome data. We fit a range of molecular clock models and assessed their statistical performance. Our analyses indicate that the emergence of VOCs is driven by an episodic increase in the substitution rate of around 4-fold the background phylogenetic rate estimate that may have lasted several weeks or months. These results underscore the importance of monitoring the molecular evolution of the virus as a means of understanding the circumstances under which VOCs may emerge.