Broad-Scale Analysis Contradicts the Theory That Generation Time Affects Molecular Evolutionary Rates in Plants

Abstract Several studies of plant taxa have concluded that generation time, including annual/perennial life history, may explain molecular evolutionary rate variation in selectively neutral DNA. Unlike in animals, there is little theoretical basis for why generation-time effects would exist in plants. Furthermore, previous reports fail to establish the generality of a generation-time effect in plants because of the small size of the datasets, a large proportion of which compared very widely divergent taxa differing in many characteristics other than generation time. Using 24 phylogenetically independent species pairs, each containing a species with an annual and a species with a perennial life history, and nine species pairs, each containing a tree species with a short and a long minimum generation time, we found no evidence that generation time is related to molecular evolutionary rate variation of the nuclear 18S ITS1 and ITS2 regions. This analysis strongly contradicts the growing belief that evolutionary rates are affected by generation time in plants. Possible reasons for the absence of generation-time effects are discussed, including an evaluation of the cell-division theory.     

Rates of molecular evolution in bacteria are relatively constant despite spore dormancy

Rates of molecular evolution are known to vary considerably among lineages, partially due to differences in life-history traits such as generation time. The generation-time effect has been well documented in some eukaryotes, but its prevalence in prokaryotes is unknown. "Because many species of Firmicute bacteria spend long periods of time as metabolically dormant spores, which could result in fewer DNA substitutions per unit time, they present an excellent system for testing predictions of the molecular clock hypothesis." To test whether spore-forming bacteria evolve more slowly than their non-spore-forming relatives, I used phylogenetic methods to determine if there were differences in rates of amino acid substitution between spore-forming and non-spore-forming lineages of Firmicute bacteria. Although rates of evolution do vary among lineages, I find no evidence for an effect of spore-formation on evolutionary rate and, furthermore, evolutionary rates are similar to those calculated for enteric bacteria. These results support the notion that variation in generation time does not affect evolutionary rates in bacterial lineages.     

Evolutionary Rates in Veronica L. (Plantaginaceae): Disentangling the Influence of Life History and Breeding System

The evolutionary rate at which DNA sequences evolve is known to differ between different groups of organisms. However, the reasons for these different rates are seldom known. Among plants, the generation-time hypothesis, which states that organisms that reproduce faster also have more DNA substitutions per time, has gained most popularity. We evaluate the generation-time hypothesis using 131 DNA sequences from the plastid trnLF region and the nuclear ribosomal ITS region of the genus Veronica (Plantaginaceae). We also examine the alternative hypothesis that a higher substitution rate is correlated with selfing breeding system. Selfing is associated with annual life history in many organisms and may thus often be the underlying reason for observed correlations of annual life history with other characters. We provide evidence that annual life history is more likely to be the responsible factor for higher substitution rates in Veronica than a selfing breeding system. Nevertheless, the way in which annual life history may influence substitution rate in detail remains unknown, and some possibilities are discussed.     

A genomic update on clostridial phylogeny: Gram‐negative spore formers and other misplaced clostridia

The class Clostridia in the phylum Firmicutes (formerly low‐G+C Gram‐positive bacteria) includes diverse bacteria of medical, environmental and biotechnological importance. The Selenomonas‐Megasphaera‐Sporomusa branch, which unifies members of the Firmicutes with Gram‐negative‐type cell envelopes, was recently moved from Clostridia to a separate class Negativicutes. However, draft genome sequences of the spore‐forming members of the Negativicutes revealed typically clostridial sets of sporulation genes. To address this and other questions in clostridial phylogeny, we have compared a phylogenetic tree for a concatenated set of 50 widespread ribosomal proteins with the trees for beta subunits of the RNA polymerase (RpoB) and DNA gyrase (GyrB) and with the 16S rRNA‐based phylogeny. The results obtained by these methods showed remarkable consistency, suggesting that they reflect the true evolutionary history of these bacteria. These data put the Selenomonas‐Megasphaera‐Sporomusa group back within the Clostridia. They also support placement of Clostridium difficile and its close relatives within the family Peptostreptococcaceae; we suggest resolving the long‐standing naming conundrum by renaming it Peptoclostridium difficile. These data also indicate the existence of a group of cellulolytic clostridia that belong to the family Ruminococcaceae. As a tentative solution to resolve the current taxonomical problems, we propose assigning 78 validly described Clostridium species that clearly fall outside the family Clostridiaceae to six new genera: Peptoclostridium, Lachnoclostridium, Ruminiclostridium, Erysipelatoclostridium, Gottschalkia and Tyzzerella. This work reaffirms that 16S rRNA and ribosomal protein sequences are better indicators of evolutionary proximity than phenotypic traits, even such key ones as the structure of the cell envelope and Gram‐staining pattern.     

ABRUPT DECELERATION OF MOLECULAR EVOLUTION LINKED TO THE ORIGIN OF ARBORESCENCE IN FERNS

Molecular rate heterogeneity, whereby rates of molecular evolution vary among groups of organisms, is a well‐documented phenomenon. Nonetheless, its causes are poorly understood. For animals, generation time is frequently cited because longer‐lived species tend to have slower rates of molecular evolution than their shorter‐lived counterparts. Although a similar pattern has been uncovered in flowering plants, using proxies such as growth form, the underlying process has remained elusive. Here, we find a deceleration of molecular evolutionary rate to be coupled with the origin of arborescence in ferns. Phylogenetic branch lengths within the “tree fern” clade are considerably shorter than those of closely related lineages, and our analyses demonstrate that this is due to a significant difference in molecular evolutionary rate. Reconstructions reveal that an abrupt rate deceleration coincided with the evolution of the long‐lived tree‐like habit at the base of the tree fern clade. This suggests that a generation time effect may well be ubiquitous across the green tree of life, and that the search for a responsible mechanism must focus on characteristics shared by all vascular plants. Discriminating among the possibilities will require contributions from various biological disciplines, but will be necessary for a full appreciation of molecular evolution.     

Molecular phylogenetic analysis of actin genic regions fromAchlya bisexualis (Oomycota) andCostaria costata (Chromophyta)

Summary Actin genic regions were isolated and characterized from the heterokont-flagellated protists,Achlya bisexualis (Oomycota) andCostaria costata (Chromophyta). Restriction enzyme and cloning experiments suggested that the genes are present in a single copy and sequence determinations revealed the existence of two introns in theC. costata actin genic region. Phylogenetic analyses of actin genic regions using distance matrix and maximum parsimony methods confirmed the close evolutionary relationship ofA. bisexualis andC. costata suggested by ribosomal DNA (rDNA) sequence comparisons and reproductive cell ultrastructure. The higher fungi, green plants, and animals were seen as monophyletic groups; however, a precise order of branching for these assemblages could not be determined. Phylogenetic frameworks inferred from comparisons of rRNAs were used to assess rates of evolution in actin genic regions of diverse eukaryotes. Actin genic regions had nonuniform rates of nucleotide substitution in different lineages. Comparison of rates of actin and rDNA sequence divergence indicated that actin genic regions evolve 2.0 and 5.3 times faster in higher fungi and flowering plants, respectively, than their rDNA sequences. Conversely, animal actins evolve at approximately one-fifth the rate of their rDNA sequences.     

Decoupled genomic elements and the evolution of partner quality in nitrogen‐fixing rhizobia

Understanding how mutualisms evolve in response to a changing environment will be critical for predicting the long‐term impacts of global changes, such as increased N (nitrogen) deposition. Bacterial mutualists in particular might evolve quickly, thanks to short generation times and the potential for independent evolution of plasmids through recombination and/or HGT (horizontal gene transfer). In a previous work using the legume/rhizobia mutualism, we demonstrated that long‐term nitrogen fertilization caused the evolution of less‐mutualistic rhizobia. Here, we use our 63 previously isolated rhizobium strains in comparative phylogenetic and quantitative genetic analyses to determine the degree to which variation in partner quality is attributable to phylogenetic relationships among strains versus recent genetic changes in response to N fertilization. We find evidence of distinct evolutionary relationships between chromosomal and pSym genes, and broad similarity between pSym genes. We also find that nifD has a unique evolutionary history that explains much of the variation in partner quality, and suggest MoFe subunit interaction sites in the evolution of less‐mutualistic rhizobia. These results provide insight into the mechanisms behind the evolutionary response of rhizobia to long‐term N fertilization, and we discuss the implications of our results for the evolution of the mutualism.     

Evolution of Escherichia coli in different carbon environments for 2,000 generations

Cellular energetics is thought to have played a key role in dictating all major evolutionary transitions in the history of life on Earth. However, how exactly cellular energetics and metabolism come together to shape evolutionary paths is not well understood. In particular, when an organism is evolved in different energy environments, what are the phenomenological differences in the chosen evolutionary trajectories, is a question that is not well understood. In this context, starting from an Escherichia coli K‐12 strain, we evolve the bacterium in five different carbon environments—glucose, arabinose, xylose, rhamnose and a mixture of these four sugars (in a predefined ratio) for approximately 2,000 generations. At the end of the adaptation period, we quantify and compare the growth dynamics of the strains in a variety of environments. The evolved strains show no specialized adaptation towards growth in the carbon medium in which they were evolved. Rather, in all environments, the evolved strains exhibited a reduced lag phase and an increased growth rate. Sequencing results reveal that these dynamical properties are not introduced via mutations in the precise loci associated with utilization of the sugar in which the bacterium evolved. These phenotypic changes are rather likely introduced via mutations elsewhere on the genome. Data from our experiments indicate that evolution in a defined environment does not alter hierarchy in mixed‐sugar utilization in bacteria.     

Competition between Moina macrocopa and Ceriodaphnia dubia: a Life Table Demography Study

We analyzed the effect of competition on survival‐ and reproduction‐related parameters under two (0.5 · 10⁶ and 1.5 · 10⁶ cells ml^—1 of Chlorella) food densities between Moina macrocopa and Ceriodaphnia dubia using the life table demography approach. In general, regardless of the presence of Moina, the average lifespan and generation time were higher for C. dubia. However, rate of population increase (r) and values of reproductive effort were higher for M. macrocopa. The net reproductive values of C. dubia grown alone varied from 7.6 to 36.6 offspring female^—1 lifetime^—1, depending on Chlorella density. Corresponding values for Moina varied little (16.4 and 17.4 offspring female^—1 lifetime^—1). Values of gross reproductive rate, net reproductive rate and the generation time of both cladoceran species were higher in mixed cultures than when grown alone. The negative effect of competition was evident for both the cladoceran species, not in their reproductive rates, but in their generation times, and thus population growth rates. The role of competition and facilitation occurring between the two species was discussed in relation to different algal food levels.     

Can plants evolve tolerance mechanisms to heterospecific pollen effects? An experimental test of the adaptive potential in Clarkia species

Flowering plants do not occur alone and often grow in mixed‐species communities where pollinator sharing is high and interactions via pollinators can occur at pre‐ and post‐pollination stages. While the causes and consequences of pre‐pollination interactions have been well studied little is known about post‐pollination interactions via heterospecific pollen (HP) receipt, and even less about the evolutionary implications of these interactions. In particular, the degree to which plants can evolve tolerance mechanisms to the negative effects of HP receipt has received little attention. Here, we aim to fill this gap in our understanding of post‐pollination interactions by experimentally testing whether two co‐flowering Clarkia species can evolve HP tolerance, and whether tolerance to specific HP ‘genotypes’ (fine‐scale local adaptation to HP) occurs. We find that Clarkia species vary in their tolerance to HP effects. Furthermore, conspecific pollen performance and the magnitude of HP effects were related to the recipient's history of exposure to HP in C. xantiana but not in C. speciosa. Specifically, better conspecific pollen performance and smaller HP effects were observed in populations of C. xantiana plants with previous exposure to HP compared to populations without such exposure. These results suggest that plants may have the potential to evolve tolerance mechanisms to HP effects but that these may occur not from the female (stigma, style) but from the male (pollen) perspective, a possibility that is often overlooked. We find no evidence for fine‐scale local adaptation to HP receipt. Studies that evaluate the adaptive potential of plants to the negative effects of HP receipt are an important first step in understanding the evolutionary consequences of plant–plant post‐pollination interactions. Such knowledge is in turn crucial for deciphering the role of plant–pollinator interactions in driving floral evolution and the composition of co‐flowering communities.     

Impact of non‐pathogenic bacteria on insect disease resistance: importance of ecological context

1. The aerial surface of plants is a habitat for large and diverse microbial communities; termed the phyllosphere. These microbes are unavoidably consumed by herbivores, and while the entomopathogens are well studied, the impact of non‐pathogenic bacteria on herbivore life history is less clear. 2. Previous work has suggested that consumption of non‐entomopathogenic bacteria induces a costly immune response that might decrease the risk of infection. However, we hypothesised that insect herbivores should be selective in how they respond to commonly encountered non‐pathogenic bacteria on their host plants to avoid unnecessary and costly immune responses. 3. An ecologically realistic scenario was used in which we fed cabbage looper, Trichoplusia ni Hübner, larvae on cabbage or cucumber leaves treated with the common non‐entomopathogenic phyllosphere bacteria, Pseudomonas fluorescens and P. syringae. Their constitutive immunity and resistance to a pathogenic bacterium (Bacillus thuringiensis; Bt) and a baculovirus (T. ni single nucleopolyhedrovirus) were then examined. 4. While feeding on bacteria‐treated leaves reduced the growth rate and condition of T. ni, there was no effect on immunity (haemolymph antibacterial and phenoloxidase activities and haemocyte numbers). Phyllosphere bacteria weakly affected the resistance of T. ni to Bt but the direction of this effect was concentration dependent; resistance to the virus was unaffected. Host plant had an impact, with cucumber‐fed larvae being more susceptible to Bt. 5. The lack of evidence for a costly immune response to non‐entomopathogenic bacteria suggests that T. ni are probably adapted to consuming common phyllosphere bacteria, and highlights the importance of the evolutionary history of participants in multi‐trophic interactions.     

Revealing hidden evolutionary capacity to cope with global change

The extent to which global change will impact the long‐term persistence of species depends on their evolutionary potential to adapt to future conditions. While the number of studies that estimate the standing levels of adaptive genetic variation in populations under predicted global change scenarios is growing all the time, few studies have considered multiple environments simultaneously and even fewer have considered evolutionary potential in multivariate context. Because conditions will not be constant, adaptation to climate change is fundamentally a multivariate process so viewing genetic variances and covariances over multivariate space will always be more informative than relying on bivariate genetic correlations between traits. A multivariate approach to understanding the evolutionary capacity to cope with global change is necessary to avoid misestimating adaptive genetic variation in the dimensions in which selection will act. We assessed the evolutionary capacity of the larval stage of the marine polychaete Galeolaria caespitosa to adapt to warmer water temperatures. Galeolaria is an important habitat‐forming species in Australia, and its earlier life‐history stages tend to be more susceptible to stress. We used a powerful quantitative genetics design that assessed the impacts of three temperatures on subsequent survival across over 30 000 embryos across 204 unique families. We found adaptive genetic variation in the two cooler temperatures in our study, but none in the warmest temperature. Based on these results, we would have concluded that this species has very little capacity to evolve to the warmest temperature. However, when we explored genetic variation in multivariate space, we found evidence that larval survival has the potential to evolve even in the warmest temperatures via correlated responses to selection across thermal environments. Future studies should take a multivariate approach to estimating evolutionary capacity to cope with global change lest they misestimate a species’ true adaptive potential.     

Toward a theory for diversity gradients: the abundance–adaptation hypothesis

The abundance–adaptation hypothesis argues that taxa with more individuals and faster generation times will have more evolutionary ‘experiments’ allowing expansion into, and diversification within, novel habitats. Thus, as older taxa have produced more individuals over time, and smaller taxa have higher population sizes and faster generation times, the Latitudinal Diversity Gradients (LDGs) of these clades should show shallower slopes. We describe the LDGs for archaea, bacteria, fungi, invertebrates and trees from six North American forests. For three focal groups – bacteria, ants, and trees – older taxa had shallower LDG slopes than the more recent, terminal taxa. Across 12 orders of magnitude of body mass, LDG slopes were steeper in larger taxa. The slopes of LDGs vary systematically with body size and clade age, underscoring the non‐canonical nature of LDGs. The steepest LDG slopes were found for the largest organisms while the smallest, from bacteria to small litter‐soil invertebrates, have shallower‐ to zero‐slope LDGs. If tropical niche conservatism is the failure of clades to adapt to, and diversify in temperate habitats, then the steep LDGs of chordates and plants likely arise from the decreased ability of clades with large individuals to adapt to the multiple challenges of extra‐tropical life.     

Establishing a community‐wide DNA barcode library as a new tool for arctic research

DNA sequences offer powerful tools for describing the members and interactions of natural communities. In this study, we establish the to‐date most comprehensive library of DNA barcodes for a terrestrial site, including all known macroscopic animals and vascular plants of an intensively studied area of the High Arctic, the Zackenberg Valley in Northeast Greenland. To demonstrate its utility, we apply the library to identify nearly 20 000 arthropod individuals from two Malaise traps, each operated for two summers. Drawing on this material, we estimate the coverage of previous morphology‐based species inventories, derive a snapshot of faunal turnover in space and time and describe the abundance and phenology of species in the rapidly changing arctic environment. Overall, 403 terrestrial animal and 160 vascular plant species were recorded by morphology‐based techniques. DNA barcodes (CO1) offered high resolution in discriminating among the local animal taxa, with 92% of morphologically distinguishable taxa assigned to unique Barcode Index Numbers (BINs) and 93% to monophyletic clusters. For vascular plants, resolution was lower, with 54% of species forming monophyletic clusters based on barcode regions rbcLa and ITS2. Malaise catches revealed 122 BINs not detected by previous sampling and DNA barcoding. The insect community was dominated by a few highly abundant taxa. Even closely related taxa differed in phenology, emphasizing the need for species‐level resolution when describing ongoing shifts in arctic communities and ecosystems. The DNA barcode library now established for Zackenberg offers new scope for such explorations, and for the detailed dissection of interspecific interactions throughout the community.     

Between semelparity and iteroparity: Empirical evidence for a continuum of modes of parity

The number of times an organism reproduces (i.e., its mode of parity) is a fundamental life‐history character, and evolutionary and ecological models that compare the relative fitnesses of different modes of parity are common in life‐history theory and theoretical biology. Despite the success of mathematical models designed to compare intrinsic rates of increase (i.e., density‐independent growth rates) between annual‐semelparous and perennial‐iteroparous reproductive schedules, there is widespread evidence that variation in reproductive allocation among semelparous and iteroparous organisms alike is continuous. This study reviews the ecological and molecular evidence for the continuity and plasticity of modes of parity—that is, the idea that annual‐semelparous and perennial‐iteroparous life histories are better understood as endpoints along a continuum of possible strategies. I conclude that parity should be understood as a continuum of different modes of parity, which differ by the degree to which they disperse or concentrate reproductive effort in time. I further argue that there are three main implications of this conclusion: (1) that seasonality should not be conflated with parity; (2) that mathematical models purporting to explain the general evolution of semelparous life histories from iteroparous ones (or vice versa) should not assume that organisms can only display either an annual‐semelparous life history or a perennial‐iteroparous one; and (3) that evolutionary ecologists should base explanations of how different life‐history strategies evolve on the physiological or molecular basis of traits underlying different modes of parity.     

The unexpected mating system of the androdioecious barnacle Chelonibia testudinaria (Linnaeus 1758)

Androdioecy was first described by Darwin in his seminal work on barnacle diversity; he identified males and hermaphrodites in the same reproductive population. Today, we realize that many androdioecious plants and animals share astonishing similarities, particularly with regard to their evolutionary history and mating system. Notably, these species were ancestrally dioecious, and their mating system has the following characteristics: hermaphrodites self‐fertilize frequently, males are more successful in large mating groups, and males have a mating advantage. A male mating advantage makes androdioecy more likely to persist over evolutionary times. Androdioecious barnacles, however, appear to persist as an outlier with a different evolutionary trajectory: they originate from hermaphroditic species. Although sexual systems of androdioecious barnacles are known, no information on the mating system of androdioecious barnacles is available. This study assessed the mating system of the androdioecious barnacle Chelonibia testudinaria. In contrast to other androdioecious species, C. testudinaria does not self‐fertilize, males do not have a mating advantage over hermaphrodites, and the average mating group is quite small, averaging only three individuals. Mating success is increased by proximity to the mate and penis length. Taken together, the mating system of C. testudinaria is unusual in comparison with other androdioecious plants and animals, and the lack of a male mating advantage suggests that the mating system alone does not provide an explanation for the maintenance of androdioecy in this species. Instead, we propose that sex‐specific life history equalizes male and hermaphroditic overall fitness.     

Latitudinal clines in alternative life histories in a geometrid moth

The relative roles of genetic differentiation and developmental plasticity in generating latitudinal gradients in life histories remain insufficiently understood. In particular, this applies to determination of voltinism (annual number of generations) in short‐lived ectotherms, and the associated trait values. We studied different components of variation in development of Chiasmia clathrata (Lepidoptera: Geometridae) larvae that originated from populations expressing univoltine, partially bivoltine or bivoltine phenology along a latitudinal gradient of season length. Indicative of population‐level genetic differentiation, larval period became longer while growth rate decreased with increasing season length within a particular phenology, but saw‐tooth clines emerged across the phenologies. Indicative of phenotypic plasticity, individuals that developed directly into reproductive adults had shorter development times and higher growth rates than those entering diapause. The most marked differences between the alternative developmental pathways were found in the bivoltine region suggesting that the adaptive correlates of the direct development evolve if exposed to selection. Pupal mass followed a complex cline without clear reference to the shift in voltinism or developmental pathway probably due to varying interplay between the responses in development time and growth rate. The results highlight the multidimensionality of evolutionary trajectories of life‐history traits, which either facilitate or constrain the evolution of integrated traits in alternative phenotypes.     

Anther‐smut fungi from more contaminated sites in Chernobyl show lower infection ability and lower viability following experimental irradiation

The long‐term contamination that followed the nuclear disaster at Chernobyl provides a case study for the effects of chronic ionizing radiation on living organisms and on their ability to tolerate or evolve resistance to such radiation. Previously, we studied the fertility and viability of early developmental stages of a castrating plant pathogen, the anther‐smut fungus Microbotryum lychnidis‐dioicae, isolated from field sites varying over 700‐fold in degree of radioactive contamination. Neither the budding rate of haploid spores following meiosis nor the karyotype structure varied with increasing radiation levels at sampling sites. Here, we assessed the ability of the same M. lychnidis‐dioicae strains to perform their whole life cycle, up to the production of symptoms in the plants, that is, the development of anthers full of fungal spores; we also assessed their viability under experimental radiation. Fungal strains from more contaminated sites had no lower spore numbers in anthers or viability, but infected host plants less well, indicating lower overall fitness due to radioactivity exposure. These findings improve our understanding of the previous field data, in which the anther‐smut disease prevalence on Silene latifolia plants caused by M. lychnidis‐dioicae was lower at more contaminated sites. Although the fungus showed relatively high resistance to experimental radiation, we found no evidence that increased resistance to radiation has evolved in populations from contaminated sites. Fungal strains from more contaminated sites even tolerated or repaired damage from a brief acute exposure to γ radiation less well than those from non‐ or less contaminated sites. Our results more generally concur with previous studies in showing that the fitness of living organisms is affected by radiation after nuclear disasters, but that they do not rapidly evolve higher tolerance.