Estimation of the Amount of DNA Polymorphism When the Neutral Mutation Rate Varies among Sites

Knowing the amount of DNA polymorphism is essential to understand the mechanism of maintaining DNA polymorphism in a natural population. The amount of DNA polymorphism can be measured by the average number of nucleotide differences per site (π), the proportion of segregating (polymorphic) site (s) and the minimum number of mutations per site (s*). Since the latter two quantities depend on the sample size, θ is often used as a measure of the amount of DNA polymorphism, where θ = 4Nμ, N is the effective population size and μ is the neutral mutation rate per site per generation. It is known that θ estimated from π, s and s* under the infinite site model can be biased when the mutation rate varies among sites. We have therefore developed new methods for estimating θ under the finite site model. Using computer simulations, it has been shown that the new methods give almost unbiased estimates even when the mutation rate varies among sites substantially. Furthermore, we have also developed new statistics for testing neutrality by modifying Tajima's D statistic. Computer simulations suggest that the new test statistics can be used even when the mutation rate varies among sites.     

Influence of nutrients and currents on the genomic composition of microbes across an upwelling mosaic

Metagenomic data sets were generated from samples collected along a coastal to open ocean transect between Southern California Bight and California Current waters during a seasonal upwelling event, providing an opportunity to examine the impact of episodic pulses of cold nutrient-rich water into surface ocean microbial communities. The data set consists of ∼5.8 million predicted proteins across seven sites, from three different size classes: 0.1–0.8, 0.8–3.0 and 3.0–200.0 μm. Taxonomic and metabolic analyses suggest that sequences from the 0.1–0.8 μm size class correlated with their position along the upwelling mosaic. However, taxonomic profiles of bacteria from the larger size classes (0.8–200 μm) were less constrained by habitat and characterized by an increase in Cyanobacteria, Bacteroidetes, Flavobacteria and double-stranded DNA viral sequences. Functional annotation of transmembrane proteins indicate that sites comprised of organisms with small genomes have an enrichment of transporters with substrate specificities for amino acids, iron and cadmium, whereas organisms with larger genomes have a higher percentage of transporters for ammonium and potassium. Eukaryotic-type glutamine synthetase (GS) II proteins were identified and taxonomically classified as viral, most closely related to the GSII in Mimivirus, suggesting that marine Mimivirus-like particles may have played a role in the transfer of GSII gene functions. Additionally, a Planctomycete bloom was sampled from one upwelling site providing a rare opportunity to assess the genomic composition of a marine Planctomycete population. The significant correlations observed between genomic properties, community structure and nutrient availability provide insights into habitat-driven dynamics among oligotrophic versus upwelled marine waters adjoining each other spatially.     

Coalescent Theory for a Partially Selfing Population

A coalescent theory for a sample of DNA sequences from a partially selfing diploid population and an algorithm for simulating such samples are developed in this article. Approximate formulas are given for the expectation and the variance of the number of segregating sites in a sample of k sequences from n individuals. Several new estimators of the important parameters θ = 4Nμ and the selfing rate s, where N and μ are, respectively, the effective population size and the mutation rate per sequence per generation, are proposed and their sampling properties are studied.     

Estimating Effective Population Size and Mutation Rate from Sequence Data Using Metropolis-Hastings Sampling

We present a new way to make a maximum likelihood estimate of the parameter 4N(e)μ (effective population size times mutation rate per site, or θ) based on a population sample of molecular sequences. We use a Metropolis-Hastings Markov chain Monte Carlo method to sample genealogies in proportion to the product of their likelihood with respect to the data and their prior probability with respect to a coalescent distribution. A specific value of θ must be chosen to generate the coalescent distribution, but the resulting trees can be used to evaluate the likelihood at other values of θ, generating a likelihood curve. This procedure concentrates sampling on those genealogies that contribute most of the likelihood, allowing estimation of meaningful likelihood curves based on relatively small samples. The method can potentially be extended to cases involving varying population size, recombination, and migration.     

Bottleneck Effect on Evolutionary Rate in the Nearly Neutral Mutation Model

Variances of evolutionary rates among lineages in some proteins are larger than those expected from simple Poisson processes. This phenomenon is called overdispersion of the molecular clock. If population size N is constant, the overdispersion is observed only in a limited range of 2Nσ under the nearly neutral mutation model, where σ represents the standard deviation of selection coefficients of new mutants. In this paper, we investigated effects of changing population size on the evolutionary rate by computer simulations assuming the nearly neutral mutation model. The size was changed cyclically between two numbers, N(1) and N(2) (N(1) > N(2)), in the simulations. The overdispersion is observed if 2N(2)σ is less than two and the state of reduced size (bottleneck state) continues for more than ~0.1/u generations, where u is the mutation rate. The overdispersion results mainly because the average fitnesses of only a portion of populations go down when the population size is reduced and only in these populations subsequent advantageous substitutions occur after the population size becomes large. Since the fitness reduction after the bottleneck is stochastic, acceleration of the evolutionary rate does not necessarily occur uniformly among loci. From these results, we argue that the nearly neutral mutation model is a candidate mechanism to explain the overdispersed molecular clock.     

A Fast Estimate for the Population Recombination Rate Based on Regression

Recombination is a fundamental evolutionary force. Therefore the population recombination rate ρ plays an important role in the analysis of population genetic data; however, it is notoriously difficult to estimate. This difficulty applies both to the accuracy of commonly used estimates and to the computational efforts required to obtain them. Some particularly popular methods are based on approximations to the likelihood. They require considerably less computational efforts than the full-likelihood method with not much less accuracy. Nevertheless, the computation of these approximate estimates can still be very time consuming, in particular when the sample size is large. Although auxiliary quantities for composite likelihood estimates can be computed in advance and stored in tables, these tables need to be recomputed if either the sample size or the mutation rate θ changes. Here we introduce a new method based on regression combined with boosting as a model selection technique. For large samples, it requires much less computational effort than other approximate methods, while providing similar levels of accuracy. Notably, for a sample of hundreds or thousands of individuals, the estimate of ρ using regression can be obtained on a single personal computer within a couple of minutes while other methods may need a couple of days or months (or even years). When the sample size is smaller (n ≤ 50), our new method remains computational efficient but produces biased estimates. We expect the new estimates to be helpful when analyzing large samples and/or many loci with possibly different mutation rates.     

Deficient SUMO Attachment to Flp Recombinase Leads to Homologous Recombination-dependent Hyperamplification of the Yeast 2 μm Circle Plasmid

Many Saccharomyces cerevisiae mutants defective in the SUMO pathway accumulate elevated levels of the native 2 μm circle plasmid (2 μm). Here we show that accumulation of 2 μm in the SUMO pathway mutants siz1Δ siz2Δ, slx5Δ, and slx8Δ is associated with formation of an aberrant high-molecular-weight (HMW) form of 2 μm. Characterization of this species from siz1Δ siz2Δ showed that it contains tandem copies of the 2 μm sequence as well as single-stranded DNA. Accumulation of this species requires both the 2 μm–encoded Flp recombinase and the cellular homologous recombination repair (HRR) pathway. Importantly, reduced SUMO attachment to Flp is sufficient to induce formation of this species. Our data suggest a model in which Flp that cannot be sumoylated causes DNA damage, whose repair via HRR produces an intermediate that generates tandem copies of the 2 μm sequence. This intermediate may be a rolling circle formed via break-induced replication (BIR), because mutants defective in BIR contain reduced levels of the HMW form. This work also illustrates the importance of using cir° strains when studying mutants that affect the yeast SUMO pathway, to avoid confusing direct functions of the SUMO pathway with secondary effects of 2 μm amplification.     

Evolutionarily stable mutation rate in a periodically changing environment

Evolution of mutation rate controlled by a neutral modifier is studied for a locus with two alleles under temporally fluctuating selection pressure. A general formula is derived to calculate the evolutionarily stable mutation rate μ(ess) in an infinitely large haploid population, and following results are obtained. (I) For any fluctuation, periodic or random: (1) if the recombination rate r per generation between the modifier and the main locus is 0, μ(ess) is the same as the optimal mutation rate μ(op) which maximizes the long-term geometric average of population fitness; and (2) for any r, if the strength s of selection per generation is very large, μ(ess) is equal to the reciprocal of the average number T of generations (duration time) during which one allele is persistently favored than the other. (II) For a periodic fluctuation in the limit of small s and r, μ(ess)T is a function of sT and rT with properties: (1) for a given sT, μ(ess)T decreases with increasing rT; (2) for sT </= 1, μ(ess)T is almost independent of sT, and depends on rT as μ(ess)T & 1.6 for rT << 1 and μ(ess)T & 6/rT for rT >> 1; and (3) for sT >/= 1, and for a given rT, μ(ess)T decreases with increasing sT to a certain minimum less than 1, and then increases to 1 asymptotically in the limit of large sT. (III) For a fluctuation consisting of multiple Fourier components (i.e., sine wave components), the component with the longest period is the most effective in determining μ(ess) (low pass filter effect). (IV) When the cost c of preventing mutation is positive, the modifier is nonneutral, and μ(ess) becomes larger than in the case of neutral modifier under the same selection pressure acting at the main locus. The value of c which makes μ(ess) equal to μ(op) of the neutral modifier case is calculated. It is argued that this value gives a critical cost such that, so long as the actual cost exceeds this value, the evolution rate at the main locus must be smaller than its mutation rate μ(ess).     

A Phylogenetic Estimator of Effective Population Size or Mutation Rate

A new estimator of the essential parameter θ = 4N(e)μ from DNA polymorphism data is developed under the neutral Wright-Fisher model without recombination and population subdivision, where N(e) is the effective population size and μ is the mutation rate per locus per generation. The new estimator has a variance only slightly larger than the minimum variance of all possible unbiased estimators of the parameter and is substantially smaller than that of any existing estimator. The high efficiency of the new estimator is achieved by making full use of phylogenetic information in a sample of DNA sequences from a population. An example of estimating θ by the new method is presented using the mitochondrial sequences from an American Indian population.     

Membrane Filtration of the Reiter Treponeme

A membrane filtration technique with commercially available membrane filters (Millipore Corp.) was effective for the removal of Reiter treponemes from liquids such as fluorescent-antibody conjugates, to which the organisms are added for adsorption. Reiter treponemes from an 8-day culture were not microscopically detectable in filtrates through membranes with a pore diameter of 0.45 μm, but treponemes were demonstrated in the filtrate by cultural methods. No organisms of the 8-day culture passed through a membrane filter having a pore size of 0.22 μm, as determined by microscopy and culture. Culture data indicated that a filter with a pore size of 0.1 μm was necessary to prevent passage of treponemes from 4-day cultures. It is recommended that a membrane filter with a pore size of 0.22 μm or smaller be used for the removal of Reiter treponemes from suspensions and that the age of the culture be considered in choosing filter pore size.     

Aerosols of Mycoplasmas, L Forms, and Bacteria: Comparison of Particle Size, Viability, and Lethality of Ultraviolet Radiation

Aerosols of microorganisms were tested for particle size by use of an Andersen sampler. Mycoplasma aerosols had an average count median diameter (CMD) of 2.1 ± 0.5 μ. Staphylococcus aureus L forms gave an average CMD of 4.6 ± 1.7 μ; the diphtheroid L form, a CMD of 3.4 ± 0.3 μ. Escherichia coli had a CMD of 5.4 ± 2.5 μ; Neisseria sicca, 3.3 ± 0.5 μ; N. meningitidis, 3.4 ± 0.2 μ. S. aureus ATCC 6538, the parent strain of the L form, yielded a CMD of 3.9 ± 1.2 μ. Candida albicans gave an average CMD of 5.9 ± 1.4 μ. All organisms tested survived aerosolizing and could be recovered in viable form for at least 1 hr. Ultraviolet radiation at 2,537 A destroyed the bacteria and mycoplasmas instantaneously, and destroyed 87% of the L forms of S. aureus, 69% of the diphtheroid L form, and 98% of the C. albicans cells. After irradiation, viable particles of the L form and C. albicans aerosols were consistently larger, indicating that clumping led to survival. Submicron size particles were found in aerosols of all species tested except C. albicans.     

Widespread Natural Occurrence of Hydroxyurea in Animals

Here we report the widespread natural occurrence of a known antibiotic and antineoplastic compound, hydroxyurea in animals from many taxonomic groups.Hydroxyurea occurs in all the organisms we have examined including invertebrates (molluscs and crustaceans), fishes from several major groups, amphibians and mammals. The species with highest concentrations was an elasmobranch (sharks, skates and rays), the little skate Leucoraja erinacea with levels up to 250 μM, high enough to have antiviral, antimicrobial and antineoplastic effects based on in vitro studies. Embryos of L. erinacea showed increasing levels of hydroxyurea with development, indicating the capacity for hydroxyurea synthesis. Certain tissues of other organisms (e.g. skin of the frog (64 μM), intestine of lobster (138 μM) gills of the surf clam (100 μM)) had levels high enough to have antiviral effects based on in vitro studies. Hydroxyurea is widely used clinically in the treatment of certain human cancers, sickle cell anemia, psoriasis, myeloproliferative diseases, and has been investigated as a potential treatment of HIV infection and its presence at high levels in tissues of elasmobranchs and other organisms suggests a novel mechanism for fighting disease that may explain the disease resistance of some groups. In light of the known production of nitric oxide from exogenously applied hydroxyurea, endogenous hydoxyurea may play a hitherto unknown role in nitric oxide dynamics.     

Physicochemical characterization of mitochondrial DNA from soybean

Mitochondrial DNA (mtDNA) of soybean (Glycine max L.) was isolated and its buoyant density was contrasted with that of nuclear (nDNA) and chloroplast (ctDNA) DNA. Each of the three DNAs banded at a single, characteristic buoyant density when centrifuged to equilibrium in a CsCl gradient. Buoyant densities were 1.694 g/cm³ for nDNA and 1.706 g/cm³ for mtDNA. These values correspond to G-C contents of 34.7 and 46.9%, respectively. Covalently closed, circular mtDNA molecules were isolated from soybean hypocotyls by ethidium bromide-cesium chloride density gradient centrifugation. Considerable variation in mtDNA circle size was observed by electron microscopy. There were seven apparent size classes with mean lengths of 5.9 μm (class 1), 10 μm (class 2), 12.9 μm (class 3), 16.6 μm (class 4), 20.4 μm (class 5), 24.5 μm (class 6), and 29.9 μm (class 7). In addition, minicircles were observed in all preparations. Partially denatured, circular mtDNA molecules with at least one representative from six of the seven observed size classes were mapped. In class 4, there appear to be at least three distinct denaturation patterns, indicating heterogeneity within this class. It is proposed that the mitochondrial genome of soybean is distributed among the different size circular molecules, several copies of the genome are contained within these classes and that the majority of the various size molecules may be a result of recombination events between circular molecules.     

Maximum Likelihood Estimation of Population Parameters

One of the most important parameters in population genetics is θ = 4N(e)μ where N(e) is the effective population size and μ is the rate of mutation per gene per generation. We study two related problems, using the maximum likelihood method and the theory of coalescence. One problem is the potential improvement of accuracy in estimating the parameter θ over existing methods and the other is the estimation of parameter λ which is the ratio of two θ's. The minimum variances of estimates of the parameter θ are derived under two idealized situations. These minimum variances serve as the lower bounds of the variances of all possible estimates of θ in practice. We then show that Watterson's estimate of θ based on the number of segregating sites is asymptotically an optimal estimate of θ. However, for a finite sample of sequences, substantial improvement over Watterson's estimate is possible when θ is large. The maximum likelihood estimate of λ = θ(1)/θ(2) is obtained and the properties of the estimate are discussed.     

De Novo Mutation Rate Variation and Its Determinants in Chlamydomonas

De novo mutations are central for evolution, since they provide the raw material for natural selection by regenerating genetic variation. However, studying de novo mutations is challenging and is generally restricted to model species, so we have a limited understanding of the evolution of the mutation rate and spectrum between closely related species. Here, we present a mutation accumulation (MA) experiment to study de novo mutation in the unicellular green alga Chlamydomonas incerta and perform comparative analyses with its closest known relative, Chlamydomonas reinhardtii. Using whole-genome sequencing data, we estimate that the median single nucleotide mutation (SNM) rate in C. incerta is μ = 7.6 × 10⁻¹⁰, and is highly variable between MA lines, ranging from μ = 0.35 × 10⁻¹⁰ to μ = 131.7 × 10⁻¹⁰. The SNM rate is strongly positively correlated with the mutation rate for insertions and deletions between lines (r > 0.97). We infer that the genomic factors associated with variation in the mutation rate are similar to those in C. reinhardtii, allowing for cross-prediction between species. Among these genomic factors, sequence context and complexity are more important than GC content. With the exception of a remarkably high C→T bias, the SNM spectrum differs markedly between the two Chlamydomonas species. Our results suggest that similar genomic and biological characteristics may result in a similar mutation rate in the two species, whereas the SNM spectrum has more freedom to diverge.     

De novo metagenomic assembly reveals abundant novel major lineage of Archaea in hypersaline microbial communities

This study describes reconstruction of two highly unusual archaeal genomes by de novo metagenomic assembly of multiple, deeply sequenced libraries from surface waters of Lake Tyrrell (LT), a hypersaline lake in NW Victoria, Australia. Lineage-specific probes were designed using the assembled genomes to visualize these novel archaea, which were highly abundant in the 0.1–0.8 μm size fraction of lake water samples. Gene content and inferred metabolic capabilities were highly dissimilar to all previously identified hypersaline microbial species. Distinctive characteristics included unique amino acid composition, absence of Gvp gas vesicle proteins, atypical archaeal metabolic pathways and unusually small cell size (approximately 0.6 μm diameter). Multi-locus phylogenetic analyses demonstrated that these organisms belong to a new major euryarchaeal lineage, distantly related to halophilic archaea of class Halobacteria. Consistent with these findings, we propose creation of a new archaeal class, provisionally named ‘Nanohaloarchaea''. In addition to their high abundance in LT surface waters, we report the prevalence of Nanohaloarchaea in other hypersaline environments worldwide. The simultaneous discovery and genome sequencing of a novel yet ubiquitous lineage of uncultivated microorganisms demonstrates that even historically well-characterized environments can reveal unexpected diversity when analyzed by metagenomics, and advances our understanding of the ecology of hypersaline environments and the evolutionary history of the archaea.     

Statistical Mechanics and the Evolution of Polygenic Quantitative Traits

The evolution of quantitative characters depends on the frequencies of the alleles involved, yet these frequencies cannot usually be measured. Previous groups have proposed an approximation to the dynamics of quantitative traits, based on an analogy with statistical mechanics. We present a modified version of that approach, which makes the analogy more precise and applies quite generally to describe the evolution of allele frequencies. We calculate explicitly how the macroscopic quantities (i.e., quantities that depend on the quantitative trait) depend on evolutionary forces, in a way that is independent of the microscopic details. We first show that the stationary distribution of allele frequencies under drift, selection, and mutation maximizes a certain measure of entropy, subject to constraints on the expectation of observable quantities. We then approximate the dynamical changes in these expectations, assuming that the distribution of allele frequencies always maximizes entropy, conditional on the expected values. When applied to directional selection on an additive trait, this gives a very good approximation to the evolution of the trait mean and the genetic variance, when the number of mutations per generation is sufficiently high (4Nμ > 1). We show how the method can be modified for small mutation rates (4Nμ → 0). We outline how this method describes epistatic interactions as, for example, with stabilizing selection.     

Recombination Accelerates Adaptation on a Large-Scale Empirical Fitness Landscape in HIV-1

Recombination has the potential to facilitate adaptation. In spite of the substantial body of theory on the impact of recombination on the evolutionary dynamics of adapting populations, empirical evidence to test these theories is still scarce. We examined the effect of recombination on adaptation on a large-scale empirical fitness landscape in HIV-1 based on in vitro fitness measurements. Our results indicate that recombination substantially increases the rate of adaptation under a wide range of parameter values for population size, mutation rate and recombination rate. The accelerating effect of recombination is stronger for intermediate mutation rates but increases in a monotonic way with the recombination rates and population sizes that we examined. We also found that both fitness effects of individual mutations and epistatic fitness interactions cause recombination to accelerate adaptation. The estimated epistasis in the adapting populations is significantly negative. Our results highlight the importance of recombination in the evolution of HIV-I.     

In vitro development of microcorms and stigma like structures in saffron (Crocus sativus L.)

Saffron is an important spice derived from the stigmas of Crocus sativus, a species belonging to the family Iridaceae. Due to its triploid nature it is sterile and is not able to set seeds, so it is propagated only by corms. The natural propagation rate of most geophytes including saffron is relatively low. An in vitro multiplication technique like micropropagation has been used for the propagation of saffron. In the present study, various explants were cultured on different nutrient media supplemented with various concentrations of plant growth regulators to standardize the best media combination for obtaining optimum response with respect to corm production and development of Stigma Like Structures (SLS). Highest response (60 %) was observed with half ovaries on G-5 media supplemented with 27 μM NAA and 44.4 μM BA followed by 55 % on LS media with 27 μM NAA and 44.4 μM BA. Maximum size (1.3 g) of microcorms were obtained from apical buds on the LS media supplemented with 21.6 μM NAA and 22.2 μM. Stigma Like Structures were developed from half ovary explants both directly and indirectly. Maximum number (120 indirectly and 20 directly) and size (5.2 cm) of SLS were obtained in G-5 medium supplemented with 27 μM NAA and 44.4 μM BA followed by 100 indirectly and 20 directly and 4.5 cm long on LS medium supplemented with 27 μM NAA and 44.4 μM BA.