Reproductive systems and evolution in vascular plants

Differences in the frequency with which offspring are produced asexually, through self-fertilization and through sexual outcrossing, are a predominant influence on the genetic structure of plant populations. Selfers and asexuals have fewer genotypes within populations than outcrossers with similar allele frequencies, and more genetic diversity in selfers and asexuals is a result of differences among populations than in sexual outcrossers. As a result of reduced levels of diversity, selfers and asexuals may be less able to respond adaptively to changing environments, and because genotypes are not mixed across family lineages, their populations may accumulate deleterious mutations more rapidly. Such differences suggest that selfing and asexual lineages may be evolutionarily short-lived and could explain why they often seem to be of recent origin. Nonetheless, the origin and maintenance of different reproductive modes must be linked to individual-level properties of survival and reproduction. Sexual outcrossers suffer from a cost of outcrossing that arises because they do not contribute to selfed or asexual progeny, whereas selfers and asexuals may contribute to outcrossed progeny. Selfing and asexual reproduction also may allow reproduction when circumstances reduce opportunities for a union of gametes produced by different individuals, a phenomenon known as reproductive assurance. Both the cost of outcrossing and reproductive assurance lead to an over-representation of selfers and asexuals in newly formed progeny, and unless sexual outcrossers are more likely to survive and reproduce, they eventually will be displaced from populations in which a selfing or asexual variant arises.     

Influences of clonality on plant sexual reproduction

Flowering plants possess an unrivaled diversity of mechanisms for achieving sexual and asexual reproduction, often simultaneously. The commonest type of asexual reproduction is clonal growth (vegetative propagation) in which parental genotypes (genets) produce vegetative modules (ramets) that are capable of independent growth, reproduction, and often dispersal. Clonal growth leads to an expansion in the size of genets and increased fitness because large floral displays increase fertility and opportunities for outcrossing. Moreover, the clonal dispersal of vegetative propagules can assist “mate finding,” particularly in aquatic plants. However, there are ecological circumstances in which functional antagonism between sexual and asexual reproductive modes can negatively affect the fitness of clonal plants. Populations of heterostylous and dioecious species have a small number of mating groups (two or three), which should occur at equal frequency in equilibrium populations. Extensive clonal growth and vegetative dispersal can disrupt the functioning of these sexual polymorphisms, resulting in biased morph ratios and populations with a single mating group, with consequences for fertility and mating. In populations in which clonal propagation predominates, mutations reducing fertility may lead to sexual dysfunction and even the loss of sex. Recent evidence suggests that somatic mutations can play a significant role in influencing fitness in clonal plants and may also help explain the occurrence of genetic diversity in sterile clonal populations. Highly polymorphic genetic markers offer outstanding opportunities for gaining novel insights into functional interactions between sexual and clonal reproduction in flowering plants.     

The agricultural pathology of ant fungus gardens

Gardens of fungus-growing ants (Formicidae: Attini) traditionally have been thought to be free of microbial parasites, with the fungal mutualist maintained in nearly pure "monocultures." We conducted extensive isolations of "alien" (nonmutualistic) fungi from ant gardens of a phylogenetically representative collection of attine ants. Contrary to the long-standing assumption that gardens are maintained free of microbial pathogens and parasites, they are in fact host to specialized parasites that are only known from attine gardens and that are found in most attine nests. These specialized garden parasites, belonging to the microfungus genus Escovopsis (Ascomycota: anamorphic Hypocreales), are horizontally transmitted between colonies. Consistent with theory of virulence evolution under this mode of pathogen transmission, Escovopsis is highly virulent and has the potential for rapid devastation of ant gardens, leading to colony mortality. The specialized parasite Escovopsis is more prevalent in gardens of the more derived ant lineages than in gardens of the more "primitive" (basal) ant lineages. Because fungal cultivars of derived attine lineages are asexual clones of apparently ancient origin whereas cultivars of primitive ant lineages were domesticated relatively recently from free-living sexual stocks, the increased virulence of pathogens associated with ancient asexual cultivars suggests an evolutionary cost to cultivar clonality, perhaps resulting from slower evolutionary rates of cultivars in the coevolutionary race with their pathogens.     

Coevolution of robustness, epistasis, and recombination favors asexual reproduction

The prevalence of sexual reproduction remains one of the most perplexing phenomena in evolutionary biology. The deterministic mutation hypothesis postulates that sexual reproduction will be advantageous under synergistic epistasis, a condition in which mutations cause a greater reduction in fitness when combined than would be expected from their individual effects. The inverse condition, antagonistic epistasis, correspondingly is predicted to favor asexual reproduction. To assess this hypothesis, we introduce a finite population evolutionary process that combines a recombination modifier formalism with a gene-regulatory network model. We demonstrate that when reproductive mode and epistasis are allowed to coevolve, asexual reproduction outcompetes sexual reproduction. In addition, no correlation is found between the level of synergistic epistasis and the fixation time of the asexual mode. However, a significant correlation is found between the level of antagonistic epistasis and asexual mode fixation time. This asymmetry can be explained by the greater reduction in fitness imposed by sexual reproduction as compared with asexual reproduction. Our findings present evidence and suggest plausible explanations that challenge both the deterministic mutation hypothesis and recent arguments asserting the importance of emergent synergistic epistasis in the maintenance of sexual reproduction.     

Population-genomic insights into the evolutionary origin and fate of obligately asexual Daphnia pulex

Despite much theoretical work, the molecular-genetic causes and evolutionary consequences of asexuality remain largely undetermined. Asexual animal species are rare, evolutionarily short-lived, and thought to suffer mutational meltdown as a result of lack of recombination. Whole-genome analysis of 11 sexual and 11 asexual genotypes of Daphnia pulex indicates that current asexual lineages are in fact very young, exhibit no signs of purifying selection against accumulating mutations, and have extremely high rates of gene conversion and deletion. The reconstruction of chromosomal haplotypes in regions containing SNP markers associated with asexuality (chromosomes VIII and IX) indicates that introgression from a sister species, Daphnia pulicaria, underlies the origin of the asexual phenotype. Silent-site divergence of the shared chromosomal haplotypes of asexuals indicates that the spread of asexuality is as recent as 1,250 y, although the origin of the meiosis-suppressing element or elements could be substantially older. In addition, using previous estimates of the gene conversion rate from Daphnia mutation accumulation lines, we are able to age each asexual lineage. Although asexual lineages originate from wide crosses that introduce elevated individual heterozygosities on clone foundation, they also appear to be constrained by the inbreeding-like effect of loss of heterozygosity that accrues as gene conversion and hemizygous deletion expose preexisting recessive deleterious alleles of asexuals, limiting their evolutionary longevity. Our study implies that the buildup of newly introduced deleterious mutations (i.e., Muller’s ratchet) may not be the dominant force imperiling nonrecombining populations of D. pulex, as previously proposed.Obligately asexual lineages are thought to suffer from elevated deleterious mutation accumulation resulting from permanent linkage at selected sites, which reduces the efficiency of selection against newly arising deleterious alleles (1–5). However, although substantial theory suggests that the diminished ability to purge new deleterious mutations will constrain the longevity of asexual populations (6–8), little is known about the genetic mechanisms responsible for a shift in reproductive mode, the consequences of asexuality for genome evolution, or the fate of asexual lineages in nature. To examine the evolution of asexual genomes, we carried out a population-genomic analysis of an asexual animal, one made interpretable by the inclusion of a parallel set of related sexual genotypes.Although most lineages of the microcrustacean Daphnia pulex are cyclically parthenogenetic, alternating between sexual and asexual phases, obligately asexual lineages have arisen polyphyletically across North America (9–11). The spread of obligate asexuality results from the proliferation of sex-limited, meiosis-suppressing genetic elements via males produced by asexual females (11). Unlike their female clone mates, these males are often capable of haploid gamete production, providing a path for transmission of the meiosis-suppressing elements to sexual populations via backcrossing. Each such event results in the production of a new asexual genotype.We investigated the genetic basis and consequences of asexuality by sequencing the entire genomes of 11 cyclically parthenogenetic isolates (hereafter sexuals) and 11 obligately asexual isolates (hereafter asexuals) from small ponds across North America (Table S1) and found that all asexual genotypes of D. pulex studied share common haplotypes of chromosomes VIII and IX, which are apparently transmitted through asexual males without recombination (unlike the remaining 10 chromosomes). Here, we describe patterns of variation across the genomes of both reproductive types, identify features unique to asexuals, and present evidence that asexual populations are quite young (≪1,000 y) but nevertheless show signs of deleterious-mutation accumulation. In addition, we show that the asexual-linked chromosome VIII/IX haplotypes arose by introgression from Daphnia pulicaria, the sister taxon to D. pulex, and that although initially harboring higher heterozygosity levels than sexuals, asexuals lose heterozygosity rapidly via deletion, gene conversion, and/or other internal homogenizing effects, which exposes preexisting, deleterious recessive alleles.     

Irritable Bowel Syndrome,Particularly the Constipation-Predominant Form,Involves an Increase in Methanobrevibacter smithii,Which Is Associated with Higher Methane Production

Background/AimsBecause Methanobrevibacter smithii produces methane, delaying gut transit, we evaluated M. smithii loads in irritable bowel syndrome (IBS) patients and healthy controls (HC).MethodsQuantitative real-time polymerase chain reaction for M. smithii was performed on the feces of 47 IBS patients (Rome III) and 30 HC. On the lactulose hydrogen breath test (LHBT, done for 25 IBS patients), a fasting methane result ≥10 ppm using 10 g of lactulose defined methane-producers.ResultsOf 47, 20 had constipation (IBS-C), 20 had diarrhea (IBS-D) and seven were not sub-typed. The M. smithii copy number was higher among IBS patients than HC (Log₁₀5.4, interquartile range [IQR; 3.2 to 6.3] vs 1.9 [0.0 to 3.4], p<0.001), particularly among IBS-C compared to IBS-D patients (Log₁₀6.1 [5.5 to 6.6] vs 3.4 [0.6 to 5.7], p=0.001); the copy number negatively correlated with the stool frequency (R=−0.420, p=0.003). The M. smithii copy number was higher among methane-producers than nonproducers (Log₁₀6.4, IQR [5.7 to 7.4] vs 4.1 [1.8 to 5.8], p=0.001). Using a receiver operating characteristic curve, the best cutoff for M. smithii among methane producers was Log₁₀6.0 (sensitivity, 64%; specificity, 86%; area under curve [AUC], 0.896). The AUC for breath methane correlated with the M. smithii copy number among methane producers (r=0.74, p=0.008). Abdominal bloating was more common among methane producers (n=9/11 [82%] vs 5/14 [36%], p=0.021).ConclusionsPatients with IBS, particularly IBS-C, had higher copy numbers of M. smithii than HC. On LHBT, breath methane levels correlated with M. smithii loads.     

Major evolutionary transitions in ant agriculture

Agriculture is a specialized form of symbiosis that is known to have evolved in only four animal groups: humans, bark beetles, termites, and ants. Here, we reconstruct the major evolutionary transitions that produced the five distinct agricultural systems of the fungus-growing ants, the most well studied of the nonhuman agriculturalists. We do so with reference to the first fossil-calibrated, multiple-gene, molecular phylogeny that incorporates the full range of taxonomic diversity within the fungus-growing ant tribe Attini. Our analyses indicate that the original form of ant agriculture, the cultivation of a diverse subset of fungal species in the tribe Leucocoprineae, evolved approximately 50 million years ago in the Neotropics, coincident with the early Eocene climatic optimum. During the past 30 million years, three known ant agricultural systems, each involving a phylogenetically distinct set of derived fungal cultivars, have separately arisen from the original agricultural system. One of these derived systems subsequently gave rise to the fifth known system of agriculture, in which a single fungal species is cultivated by leaf-cutter ants. Leaf-cutter ants evolved remarkably recently ( approximately 8-12 million years ago) to become the dominant herbivores of the New World tropics. Our analyses identify relict, extant attine ant species that occupy phylogenetic positions that are transitional between the agricultural systems. Intensive study of those species holds particular promise for clarifying the sequential accretion of ecological and behavioral characters that produced each of the major ant agricultural systems.     

Enhanced heterozygosity from male meiotic chromosome chains is superseded by hybrid female asexuality in termites

Although males are a ubiquitous feature of animals, they have been lost repeatedly in diverse lineages. The tendency for obligate asexuality to evolve is thought to be reduced in animals whose males play a critical role beyond the contribution of gametes, for example, via care of offspring or provision of nuptial gifts. To our knowledge, the evolution of obligate asexuality in such species is unknown. In some species that undergo frequent inbreeding, males are hypothesized to play a key role in maintaining genetic heterozygosity through the possession of neo-sex chromosomes, although empirical evidence for this is lacking. Because inbreeding is a key feature of the life cycle of termites, we investigated the potential role of males in promoting heterozygosity within populations through karyotyping and genome-wide single-nucleotide polymorphism analyses of the drywood termite Glyptotermes nakajimai. We showed that males possess up to 15 out of 17 of their chromosomes as sex-linked (sex and neo-sex) chromosomes and that they maintain significantly higher levels of heterozygosity than do females. Furthermore, we showed that two obligately asexual lineages of this species—representing the only known all-female termite populations—arose independently via intraspecific hybridization between sexual lineages with differing diploid chromosome numbers. Importantly, these asexual females have markedly higher heterozygosity than their conspecific males and appear to have replaced the sexual lineages in some populations. Our results indicate that asexuality has enabled females to supplant a key role of males.

Although asexual populations should have a twofold reproductive advantage over their sexual relatives (1), sexual reproduction is the rule in almost all animals and plants (2). This is probably because sexual reproduction enables gene pools to be constantly mixed, generates new combinations of genes, and facilitates adaptation to complex and heterogeneous environments (3). Nevertheless, obligately asexual lineages have evolved repeatedly in diverse animal taxa (2, 4, 5), which remains an important unsolved problem in evolutionary biology. Many biologists have approached this problem by considering the advantages of asexuality and how the disadvantages of asexuality can be circumvented (6–8). In each case, it is thought that the evolution of asexuality should be prevented when males have crucial roles in the biology and life cycle of a species or population (e.g., paternal care for offspring and nuptial gifts for females) (1, 9, 10). Indeed, to our knowledge, the evolution of obligate asexual lineages from ancestors whose males play a critical role beyond that of gamete provision is unknown.In inbred populations of some species, males potentially play a key role in maintaining heterozygosity through the possession of neo-sex chromosomes (11). Such chromosomal systems are found in some animals and plants, arising as a result of reciprocal translocations or centric fusions between sex chromosomes and autosomes (12–15). Under male heterogamety (i.e., XY = male, XX = female), it has been hypothesized that autosomes that are linked to the Y chromosome (i.e., neo-Y chromosomes) during meiosis never become homozygous by descent in the absence of crossing-over, allowing maintenance of heterozygosity (11). Therefore, neo-Y chromosomes would help lineages that undergo frequent inbreeding to reduce genetic costs of inbreeding in males. However, to our knowledge, there have been no empirical tests of this hypothesis. Furthermore, the potential role of males in maintaining heterozygosity is also expected to reduce the tendency for males to be lost through the evolution of asexuality.Termites provide an ideal model to explore the role of males in animal species, in particular those species which undergo regular inbreeding. This is because, although almost all termite species undergo outbreeding during swarms of virgin reproductives, inbreeding as a result of sibling–sibling or parent–sibling reproduction within nests appears to be a key feature of the life cycle of many species (16, 17). Nevertheless, reduced genetic heterozygosity in termites caused by inbreeding can result not only in individual-level costs (e.g., reduced fecundity) but also in colony-level costs (e.g., reduced disease resistance) (18, 19). Such inbreeding is thought to have given rise to a striking karyological feature of many termite species: the formation of chains (or rings) of several chromosomes (sex chromosomes [i.e., X and Y chromosomes] plus autosomes [i.e., neo-X and neo-Y chromosomes]) during male meiosis, whereby the Y chromosomes and some autosomes (i.e., neo-Y chromosomes) segregate together as a single linkage group to male-determining sperm (i.e., a neo-Y chromosome system) (14, 20, 21). Heterozygote advantage in the face of inbreeding has been postulated to account for the evolution of this system (11), although extensive genetic analyses examining the effects of neo-Y chromosome systems have not yet been conducted.We have recently investigated the biology of Glyptotermes nakajimai Morimoto (Isoptera: Kalotermitidae) (22), a species of drywood termite found in southern areas of the mainland of Japan, as well as islands farther south (23). We examined sex and caste ratios within colonies, sperm storage of egg-laying queens, and hatching success of unfertilized eggs. We discovered the presence of up to 25 secondary (neotenic) reproductives (i.e., offspring of primary kings and queens) in most field colonies, suggesting that inbreeding occurs in this species. Despite the presumed role of males in maintaining heterozygosity in termite populations (described above), we have discovered a number of asexual (all-female) G. nakajimai populations—an evolutionary transition from mixed-sex to all-female asexual societies (22). Although individuals from asexual and sexual populations are indistinguishable by external morphology and cuticular hydrocarbon profiles (23), previous molecular phylogenetic analyses have shown that asexual and sexual populations respectively form separate monophyletic groups (22). Notably, individuals of asexual populations have an uneven number of chromosomes (2n = 35), in contrast to those of sexual populations (2n = 34) (22). An uneven number of chromosomes in a diploid organism, in particular in females, can arise through hybridization between closely related lineages that differ in diploid chromosome number (e.g., refs. 24 and 25). Such hybrids are expected to be sterile due to chromosome pairing incompatibilities during meiosis, providing an opportunity for the evolution of asexuality (13, 26). Importantly, hybrid asexuals in other species are known to often exhibit high and fixed heterozygosity due to the combination of two different genomes (27).To investigate the evolution of asexuality in species that undergo inbreeding, we used G. nakajimai as a model species. We performed a series of analyses based on genome-wide single-nucleotide polymorphisms (SNPs) generated in representatives across the distribution of this species and examined the karyotypes of selected populations. We sought to address the following questions: 1) What is the population genetic structure of G. nakajimai, and how are sexual and asexual G. nakajimai individuals related to each other? 2) Do male G. nakajimai possess neo-sex chromosomes, and does heterozygosity vary between males, sexual females, and asexual females? 3) Did asexual G. nakajimai arise via hybridization, as predicted on the basis of chromosome number?     

Methanobrevibacter smithii Is the Predominant Methanogen in Patients with Constipation-Predominant IBS and Methane on Breath

Purpose

Among irritable bowel syndrome (IBS) patients, breath methane producers overwhelmingly have constipation predominance (C-IBS). Although the most common methanogen in humans is Methanobrevibacter smithii, incidence and type of methanogenic bacteria in C-IBS patients are unknown.

Methods

By use of a questionnaire and lactulose breath testing, subjects with Rome II C-IBS and methane (>3?ppm) were selected (n?=?9). The control group included subjects with IBS who had no breath methane (n?=?10). Presence of bacterial DNA was assessed in a stool sample of each subject by quantitative-PCR using universal 16S rDNA primer. M. smithii was quantified by use of a specific rpoB gene primer.

Results

M. smithii was detected in both methane and non-methane subjects. However, counts and relative proportion of M. smithii were significantly higher for methane-positive than for methane-negative subjects (1.8?×?10⁷?±?3.0?×?10⁷ vs 3.2?×?10⁵?±?7.6?×?10⁵?copies/g wet stool, P?<?0.001; and 7.1?±?6.3?% vs 0.24?±?0.47?%, P?=?0.02 respectively). The minimum threshold of M. smithii resulting in positive lactulose breath testing for methane was 4.2?×?10⁵?copies/g wet stool or 1.2?% of total stool bacteria. Finally, area-under-curve for breath methane correlated significantly with both absolute quantity and percentage of M. smithii in stool (R?=?0.76; P?<?0.001 and R?=?0.77; P?<?0.001 respectively).

Conclusions

M. smithii is the predominant methanogen in C-IBS patients with methane on breath testing. The number and proportion of M. smithii in stool correlate well with amount of breath methane.     

Rare gene capture in predominantly androgenetic species

The long-term persistence of completely asexual species is unexpected. Although asexuality has short-term evolutionary advantages, a lack of genetic recombination leads to the accumulation over time of deleterious mutations. The loss of individual fitness as a result of accumulated deleterious mutations is expected to lead to reduced population fitness and possible lineage extinction. Persistent lineages of asexual, all-female clones (parthenogenetic and gynogenetic species) avoid the negative effects of asexual reproduction through the production of rare males, or otherwise exhibit some degree of genetic recombination. Another form of asexuality, known as androgenesis, results in offspring that are clones of the male parent. Several species of the Asian clam genus Corbicula reproduce via androgenesis. We compared gene trees of mitochondrial and nuclear loci from multiple sexual and androgenetic species across the global distribution of Corbicula to test the hypothesis of long-term clonality of the androgenetic species. Our results indicate that low levels of genetic capture of maternal nuclear DNA from other species occur within otherwise androgenetic lineages of Corbicula. The rare capture of genetic material from other species may allow androgenetic lineages of Corbicula to mitigate the effects of deleterious mutation accumulation and increase potentially adaptive variation. Models comparing the relative advantages and disadvantages of sexual and asexual reproduction should consider the possibility of rare genetic recombination, because such events seem to be nearly ubiquitous among otherwise asexual species.     

Sexual development in the industrial workhorse Trichoderma reesei

Filamentous fungi are indispensable biotechnological tools for the production of organic chemicals, enzymes, and antibiotics. Most of the strains used for industrial applications have been—and still are—screened and improved by classical mutagenesis. Sexual crossing approaches would yield considerable advantages for research and industrial strain improvement, but interestingly, industrially applied filamentous fungal species have so far been considered to be largely asexual. This is also true for the ascomycete Trichoderma reesei (anamorph of Hypocrea jecorina), which is used for production of cellulolytic and hemicellulolytic enzymes. In this study, we report that T. reesei QM6a has a MAT1-2 mating type locus, and the identification of its respective mating type counterpart, MAT1-1, in natural isolates of H. jecorina, thus proving that this is a heterothallic species. After being considered asexual since its discovery more than 50 years ago, we were now able to induce sexual reproduction of T. reesei QM6a and obtained fertilized stromata and mature ascospores. This sexual crossing approach therefore opens up perspectives for biotechnologically important fungi. Our findings provide a tool for fast and efficient industrial strain improvement in T. reesei, thus boosting research toward economically feasible biofuel production. In addition, knowledge of MAT-loci and sexual crossing techniques will facilitate research with other Trichoderma spp. relevant for agriculture and human health.     

Does the avoidance of sexual costs increase fitness in asexual invaders?

The high prevalence of sexual reproduction is considered a paradox mainly for two reasons. First, asexuals should enjoy various growth benefits because they seemingly rid themselves of the many inefficiencies of sexual reproduction—the so-called costs of sex. Second, there seems to be no lack of asexual origins because losses of sexual reproduction have been described in almost every larger eukaryotic taxon. Current attempts to resolve this paradox concentrate on a few hypotheses that provide universal benefits that would compensate for these costs and give sexual reproduction a net advantage. However, are new asexual lineages really those powerful invaders that could quickly displace their sexual ancestors? Research on the costs of sex indicates that sex is often stabilized by highly lineage-specific mechanisms. Two main categories can be distinguished. First are beneficial traits that evolved within a particular species and became tightly associated with sex (e.g., a mating system that involves sexual selection, or a sexual diapausing stage that allows survival through harsh periods). If such traits are absent in asexuals, simple growth efficiency considerations will not capture the fitness benefits gained by skipping sexual reproduction. Second, lineage-specific factors might prevent asexuals from reaching their full potential (e.g., dependence on fertilization in sperm-dependent parthenogens). Such observations suggest that the costs of sex are highly variable and often lower than theoretical considerations suggest. This has implications for the magnitude of universal benefits required to resolve the paradox of sex.     

Small genome of the fungus Escovopsis weberi,a specialized disease agent of ant agriculture

Many microorganisms with specialized lifestyles have reduced genomes. This is best understood in beneficial bacterial symbioses, where partner fidelity facilitates loss of genes necessary for living independently. Specialized microbial pathogens may also exhibit gene loss relative to generalists. Here, we demonstrate that Escovopsis weberi, a fungal parasite of the crops of fungus-growing ants, has a reduced genome in terms of both size and gene content relative to closely related but less specialized fungi. Although primary metabolism genes have been retained, the E. weberi genome is depleted in carbohydrate active enzymes, which is consistent with reliance on a host with these functions. E. weberi has also lost genes considered necessary for sexual reproduction. Contrasting these losses, the genome encodes unique secondary metabolite biosynthesis clusters, some of which include genes that exhibit up-regulated expression during host attack. Thus, the specialized nature of the interaction between Escovopsis and ant agriculture is reflected in the parasite’s genome.The highly evolved agricultural lifestyle of leaf-cutting ants has attracted particular attention because these ants cultivate a symbiotic fungus that serves as their major food source. These ants cut leaves, preprocess them into small pieces, and feed them to the cultivated fungus (1). The capacity of the cultivated fungus to break down plant material gives ant agriculturalists access to the vast nutrient stores locked within neotropical plants (Fig. 1A) (2–5). The symbiosis between fungus-growing ants and their cultivated fungi has persisted for at least 50 million years (6).Open in a separate windowFig. 1.Escovopsis weberi, a specialized mycoparasite of the fungus-growing ant symbiosis, has a small genome compared with other Pezizomycotina fungi. (A) Both fungus-growing ants and the mycoparasite E. weberi use the ants’ cultivated fungi as their primary food source. The ability of the cultivated fungi to efficiently break down plant material gives both consumers access to the biomass of neotropical plants. (B) Size and protein-coding gene content of genomes of diverse fungi in the Pezizomycotina. Bayesian phylogeny estimated using partial amino acid alignments of three genes (Rpb1, Rpb2, ef1-α). All posterior probabilities are greater than 0.95. Phylogeny is rooted with Sacchormyces cervesiae (not shown). (C) Relationship between genome size and gene content. A list of genomes included in this panel is in SI Appendix, Table S1.Like human agriculture, ant agriculture is hampered by disease. The ants’ fungal crops are attacked and consumed by fungal parasites of the genus Escovopsis (Ascomycota, Pezizomycotina: anamorphic Hypocreales) (Fig. 1A) (7), which have evolved in association with the ants and their cultivated fungi (8). Escovopsis infection can have detrimental impacts on garden health and, consequently, on the survival of ant colonies (9, 10). Such mycoparasitism, the phenomenon whereby one fungus is parasitic on another fungus, is rare. It is most well-known for species from the genus Trichoderma, some of which are used as biocontrol agents for fungal diseases and others of which attack human-cultivated fungi (11–13). In contrast to Trichoderma species, however, Escovopsis species grow poorly in their hosts’ absence (SI Appendix, Figs. S1 and S2).Escovopsis species have never been isolated outside of fungus-growing ant colonies, and different strains of Escovopsis are capable of attacking the fungi grown by different fungus-growing ant species (8, 14, 15). The long-term, specialized evolutionary history of the association between Escovopsis and their hosts provides a unique venue to explore the consequences of host specialization on pathogen genome evolution. Here, we assemble and annotate the genome of a strain of Escovopsis weberi. Consistent with expectations under an evolutionary transition toward using a narrow host range, and similar to many other specialized, host-associated microbes (16, 17), E. weberi exhibits gene loss. Contrasting other fungal pathogens, the large genomes of which are expanded with genetic elements that influence host adaptation (18), the genome size of Escovopsis is small compared with those of its closest sequenced relatives.     

Cryptic sex and many-to-one coevolution in the fungus-growing ant symbiosis

The fungus-growing ants have long provided a spectacular example of coevolutionary integration. Their ecological success is thought to depend largely on the evolutionary alignment of reproductive interests between ants and fungi after vertical transmission and the ancient suppression of fungal sexuality. In the present study we test these assumptions and provide the first evidence of recombination in attine cultivars, contradicting widely held perceptions of obligate clonality. In addition, we document long-distance horizontal transmission of symbionts between leaf-cutter ant species on mainland Central America and South America and those endemic to Cuba, suggesting both lack of pairwise coevolutionary specificity in ant/cultivar interactions and dispersal of symbionts independent of their ant hosts. The coevolution between leaf-cutters and their fungal symbionts is thus not reciprocally pairwise. Rather, a single widespread and sexual fungal symbiont species is engaged in multiple interactions with divergent ant lineages. Strict fungal clonality and vertical transmission evidently have not played a critical role in the long-term evolutionary or ecological success of this well known mutualism.     

Population genomics reveals a possible history of backcrossing and recombination in the gynogenetic fish Poecilia formosa

Unisexual sperm-dependent vertebrates are of hybrid origins, rare, and predicted to be short-lived as a result of several challenges arising from their mode of reproduction. In particular, because of a lack of recombination, clonal species are predicted to have a low potential to respond to natural selection. However, many unisexual sperm-dependent species persist, and assessing the genetic diversity present in these species is fundamental to understanding how they avoid extinction. We used population genomic methods to assess genotypic variation within the unisexual fish Poecilia formosa. Measures of admixture and population differentiation, as well as clustering analyses, indicate that the genomes of individuals of P. formosa are admixed and intermediate between Poecilia latipinna and Poecilia mexicana, consistent with the hypothesis of their hybrid origins. Bayesian genomic cline analyses indicate that about 12% of sampled loci exhibit patterns consistent with inheritance from only one parent. The estimation of observed heterozygosity clearly suggests that P. formosa is not comprised of direct descendants of a single nonrecombining asexual F₁ hybrid individual. Additionally, the estimation of observed heterozygosity provides support for the hypothesis that the history of this unisexual species has included backcrossing with the parent species before the onset of gynogenesis. We also document high levels of variation among asexual individuals, which is attributable to recombination (historical or ongoing) and the accumulation of mutations. The high genetic variation suggests that this unisexual vertebrate has more potential to respond to natural selection than if they were frozen F₁ hybrids.The maintenance of sex presents a conundrum for evolutionary biology because the costs of sexual reproduction (cost of producing males, energy expenditure to find a mate, exposure to diseases, and segregation of alleles) appear to be immediate and substantial, whereas its benefits (facilitation of adaptations, elimination of deleterious mutations) are postponed (reviewed in ref. 1). The long-term maintenance of unisexual organisms is of interest to evolutionary biologists as well because the advantages of asexual reproduction are all immediate (no cost of producing males and, therefore, exponential population growth), but the long-term costs are substantial (accumulation of deleterious mutations and lack of genetic recombination to respond to environmental changes). Asexual vertebrate species are, therefore, predicted to be short-lived compared with sexually reproducing species (2–4). However, recent work focused on nonvertebrate species has challenged the view that recombination is absent in asexual lineages and that, therefore, those species are doomed to extinction. Asexual aphids, fungi, and microcrustaceans have all been shown to be genetically variable [aphids (5), fungi (6), Daphnia (7)] and, in some cases, mitotic recombination facilitates the spread of beneficial mutations (8). Therefore, understanding how much genetic variation is present in asexual lineages, and whether the presence of this variation and the mechanisms that facilitate it are shared among taxa, is an essential step toward understanding the evolution of sexual and asexual reproduction and, perhaps, challenging existing paradigms.Asexuality is common in many phyla (reviewed in ref. 7), but it is relatively rare in vertebrates (1). All known unisexual (all-female) vertebrates are products of hybridization events between sexually reproducing species (ref. 9 and references therein), constitute only 0.1% of extant vertebrate species (1, 9). One type of asexual reproduction found in unisexual vertebrates is gynogenesis, where females must mate with males of a closely related species (but refer to ref. 10 for exceptions), but the nonrecombinant embryos do not inherit any genetic information from the sperm donor (9). Because gynogens require sperm to initiate development of offspring, but no paternal genes are expressed, they are considered “sexual parasites” (11).The maintenance of a gynogenetic species is paradoxical because gynogens face the costs of both sexual and asexual reproduction: the cost of finding a mate, exposure to diseases, accumulation of deleterious mutations, and lack of genetic recombination to facilitate adaptation. In addition, because male sperm donors do not gain a fitness advantage from mating with gynogens, selection should favor males that avoid mating with them.Given the extensive and diverse list of challenges faced by gynogenetic species, they are predicted to be short-lived with a limited potential to respond to natural selection. Nevertheless, gynogenetic species persist, and some have origins in the distant past (12, 13). This suggests that gynogenetic species might be able to avoid or ameliorate some of the costs associated with their reproductive mode. One question that arises is how much genetic variation persists in gynogenetic species?The Amazon molly (Poecilia formosa) is an excellent system to explore this question. P. formosa is the first vertebrate recognized as asexual (11) and is a gynogenetic species that uses Poecilia mexicana (Atlantic molly), Poecilia latipinna (sailfin molly), and Poecilia latipunctata (Tamesi molly) as sexual hosts (14). Like every other known unisexual vertebrate, P. formosa is thought to be a hybrid lineage (9, 13, 15–17). P. mexicana is recognized to be the maternal species of P. formosa (13, 15–17), whereas P. latipinna (or an extinct ancestor of P. latipinna) is the putative paternal species (17). P. formosa lives in sympatry with at least one of the two parent species throughout its range from the Tampico region in Mexico to the southeastern United States (Fig. 1A). Although recent studies suggest that P. formosa is a species that consists of “frozen” F₁ hybrid clones (i.e., individuals with ancestry from P. mexicana and P. latipinna at all loci) (15–19), this result is based on limited genetic data. Additionally, it is still not clear whether P. formosa is the product of a single or multiple hybridization events, although a recent investigation supports the hypothesis of a single event possibly giving rise to several clonal lineages (19).Open in a separate windowFig. 1.Sampled populations (A) and PCA plots for the 192 individuals based on the genotype probabilities at each locus (B–D). Dark blue, north P. latipinna; blue, central P. latipinna; light blue, south P. latipinna; orange, north P. formosa sympatric with P. latipinna; red, south P. formosa sympatric with P. mexicana; light green, north P. mexicana; green, central P. mexicana; dark green, south P. mexicana.The overall objective of this research was to examine genotypic variation in P. formosa at a genomic scale. We generated thousands of DNA sequence markers to address three main questions. (i) Is P. formosa the product of hybridization between P. latipinna and P. mexicana? This is the conclusion of several previous studies (11, 13, 15–19), and, here, we attempt to confirm this result with a genome-wide survey of P. formosa and its putative parent species. (ii) Is P. formosa composed of frozen F₁ hybrid clones, or is there evidence of a more complicated history of nonclonal reproduction or recombination? Previous genetic analyses (13, 15–19), and the fact that no laboratory has been able to synthetize P. formosa from artificial hybridization experiments (15, 19–21), suggest that the genome of this species is more complicated than that of a simple F₁ hybrid. If P. formosa is not composed of clones of F₁ hybrid lineages, then, what is the genetic contribution of each parent species? (iii) How much genotypic diversity exists within P. formosa and is this genotypic diversity consistent with P. formosa having evolved from a single or multiple independently formed hybrid individuals? Quantifying genetic variation will provide some estimate of this species’ potential to respond to selection.     

Frequency-dependent selection maintains clonal diversity in an asexual organism

Asexual organisms can be genetically variable and evolve through time, yet it is not known how genetic diversity is maintained in populations. In sexual organisms, negative frequency-dependent selection plays a role in maintaining diversity at some loci, but in asexual organisms, this mechanism could provide a general explanation for persistent genetic diversity because it acts on the whole genome and not just on some polymorphisms within a genome. Using field manipulations, we show that negative frequency-dependent selection maintains clonal diversity in an asexual mite species, and we link predicted equilibrium clonal frequencies to average frequencies in space and time. Intense frequency-dependent selection is likely to be a general mechanism for persistent genetic diversity in asexual organisms.     

Plant sex and the evolution of plant defenses against herbivores

Despite the importance of plant–herbivore interactions to the ecology and evolution of terrestrial ecosystems, the evolutionary factors contributing to variation in plant defenses against herbivores remain unresolved. We used a comparative phylogenetic approach to examine a previously untested hypothesis (Recombination-Mating System Hypothesis) that posits that reduced sexual reproduction limits adaptive evolution of plant defenses against arthropod herbivores. To test this hypothesis we focused on the evening primrose family (Onagraceae), which includes both sexual and functionally asexual species. Ancestral state reconstructions on a 5-gene phylogeny of the family revealed between 18 and 21 independent transitions between sexual and asexual reproduction. Based on these analyses, we examined susceptibility to herbivores on 32 plant species representing 15 independent transitions. Generalist caterpillars consumed 32% more leaf tissue, gained 13% greater mass, and experienced 21% higher survival on functionally asexual than on sexual plant species. Survival of a generalist feeding mite was 19% higher on asexual species. In a field experiment, generalist herbivores consumed 64% more leaf tissue on asexual species. By contrast, a specialist beetle fed more on sexual than asexual species, suggesting that a tradeoff exists between the evolution of defense to generalist and specialist herbivores. Measures of putative plant defense traits indicate that both secondary compounds and physical leaf characteristics may mediate this tradeoff. These results support the Recombination-Mating System Hypothesis and suggest that variation in sexual reproduction among plant species may play an important, yet overlooked, role in shaping the macroevolution of plant defenses against arthropod herbivores.     

Clonal reproduction in fungi

Research over the past two decades shows that both recombination and clonality are likely to contribute to the reproduction of all fungi. This view of fungi is different from the historical and still commonly held view that a large fraction of fungi are exclusively clonal and that some fungi have been exclusively clonal for hundreds of millions of years. Here, we first will consider how these two historical views have changed. Then we will examine the impact on fungal research of the concept of restrained recombination [Tibayrenc M, Ayala FJ (2012) Proc Natl Acad Sci USA 109 (48):E3305–E3313]. Using animal and human pathogenic fungi, we examine extrinsic restraints on recombination associated with bottlenecks in genetic variation caused by geographic dispersal and extrinsic restraints caused by shifts in reproductive mode associated with either disease transmission or hybridization. Using species of the model yeast Saccharomyces and the model filamentous fungus Neurospora, we examine intrinsic restraints on recombination associated with mating systems that range from strictly clonal at one extreme to fully outbreeding at the other and those that lie between, including selfing and inbreeding. We also consider the effect of nomenclature on perception of reproductive mode and a means of comparing the relative impact of clonality and recombination on fungal populations. Last, we consider a recent hypothesis suggesting that fungi thought to have the most severe intrinsic constraints on recombination actually may have the fewest.