Asymmetric density-dependent competition does not contribute to the maintenance of sex in a mixed population of sexual and asexual Potamopyrgus antipodarum.

Asexual reproduction is expected to have a twofold reproductive advantage over sexual reproduction, owing to the cost of producing males in sexual subpopulations. The persistence of sexual females, thus, requires an advantage to sexual reproduction, at least periodically. Here, we tested the hypothesis that asexual females are more sensitive to limited resources. Under this idea, fluctuations in the availability of resources (per capita) could periodically favour sexual females when resources become limited. We combined sexual and asexual freshwater snails (Potamopyrgus antipodarum) together in nylon mesh enclosures at three different densities in an outdoor mesocosm. After 1 month, we counted the brood size of fertile female snails. We found that fecundity declined significantly with increasing density. However, sexual females did not produce more offspring than asexual females at any of the experimental densities. Our results, thus, suggest that the cost of sexual reproduction in P. antipodarum is not ameliorated by periods of intense resource competition.

Pre- and post-association barriers to host switching in sympatric mutualists.

Coevolution between mutualists can lead to reciprocal specialization, potentially causing barriers to host switching. Here, we conducted assays to identify pre- and post-association barriers to host switching by endosymbiotic bacteria, both within and between two sympatric nematode clades. In nature, Steinernema nematodes and Xenorhabdus bacteria form an obligate mutualism. Free-living juvenile nematodes carry Xenorhabdus in a specialized intestinal receptacle. When nematodes enter an insect, they release the bacteria into the insect hemocoel. The bacteria aid in killing the insect and facilitate nematode reproduction. Prior to dispersing from the insect, juvenile nematodes must form an association with their symbionts; the bacteria must adhere to the intestinal receptacle. We tested for pre-association barriers by comparing the effects of bacterial strains on native versus non-native nematodes via their virulence towards, nutritional support of, and ability to associate with different nematode species. We then assessed post-association barriers by measuring the relative fitness of nematodes carrying each strain of bacteria. We found evidence for both pre- and post-association barriers between nematode clades. Specifically, some bacteria were highly virulent to non-native hosts, and some nematode hosts carried fewer cells of non-native bacteria, creating pre-association barriers. In addition, reduced infection success and lower nematode reproduction were identified as post-association barriers. No barriers to symbiont switching were detected between nematode species within the same clade. Overall, our study suggests a framework that could be used to generate predictions for the evolution of barriers to host switching in this and other systems.

Parasitic manipulation or by-product of infection: an experimental approach using trematode-infected snails.

Natural selection should favour parasite genotypes that manipulate hosts in ways that enhance parasite fitness. However, it is also possible that the effects of infection are not adaptive. Here we experimentally examined the phenotypic effects of infection in a snail-trematode system. These trematodes (Atriophallophorus winterbourni) produce larval cysts within the snail's shell (Potamopyrgus antipodarum); hence the internal shell volume determines the total number of parasite cysts produced. Infected snails in the field tend to be larger than uninfected snails, suggesting the hypothesis that parasites manipulate host growth so as to increase the space available for trematode reproduction. To test the hypothesis, we exposed juvenile snails to trematode eggs. Snails were then left to grow for about one year in 800-l outdoor mesocosms. We found that uninfected males were smaller than uninfected females (sexual dimorphism). We also found that infection did not affect the shell dimensions of males. However, infected females were smaller than uninfected females. Hence, infection stunts the growth of females, and (contrary to the hypothesis) it results in a smaller internal volume for larval cysts. Finally, infected females resembled males in size and shape, suggesting the possibility that parasitic castration prevents the normal development of females. These results thus indicate that the parasite is not manipulating the growth of infected hosts so as to increase the number of larval cysts, although alternative adaptive explanations are possible.

Causation without correlation: parasite-mediated frequency-dependent selection and infection prevalence.

Parasite-mediated selection is thought to maintain host genetic diversity for resistance. We might thus expect to find a strong positive correlation between host genetic diversity and infection prevalence across natural populations. Here, we used computer simulations to examine host-parasite coevolution in 20 simi-isolated clonal populations across a broad range of values for both parasite virulence and parasite fecundity. We found that the correlation between host genetic diversity and infection prevalence can be significantly positive for intermediate values of parasite virulence and fecundity. But the correlation can also be weak and statistically non-significant, even when parasite-mediated frequency-dependent selection is the sole force maintaining host diversity. Hence correlational analyses of field populations, while useful, might underestimate the role of parasites in maintaining host diversity.

DNA Content Variation and SNP Diversity Within a Single Population of Asexual Snails.

A growing body of research suggests that many clonal populations maintain genetic diversity even without occasional sexual reproduction. The purpose of our study was to document variation in single-nucleotide polymorphism (SNP) diversity, DNA content, and pathogen susceptibility in clonal lineages of the New Zealand freshwater snail, Potamopyrgus antipodarum. We studied snails that were collected from multiple field sites around a single lake (Lake Alexandrina), as well as isofemale clonal lineages that had been isolated and maintained in the laboratory. We used the kompetitive allele specific PCR (KASP) method to genotype our samples at 46 nuclear SNP sites, and we used flow cytometry to estimate DNA content. We found high levels of SNP diversity, both in our field samples and in our clonal laboratory lines. We also found evidence of high variation in DNA content among clones, even among clones with identical genotypes across all SNP sites. Controlled pathogen exposures of the laboratory populations revealed variation in susceptibility among distinct clonal genotypes, which was independent of DNA content. Taken together, these results show high levels of diversity among asexual snails, especially for DNA content, and they suggest rapid genome evolution in asexuals.

Periodic, Parasite-Mediated Selection For and Against Sex.

Asexual lineages should rapidly replace sexual populations. Why sex then? The Red Queen hypothesis proposes that parasite-mediated selection against common host genotypes could counteract the per capita birth rate advantage of asexuals. Under the Red Queen hypothesis, fluctuations in parasite-mediated selection can drive fluctuations in the asexual population, leading to the coexistence of sexual and asexual reproduction. Does shifting selection by parasites drive fluctuations in the fitness and frequency of asexuals in nature? Combining long-term field data with mesocosm experiments, we detected a shift in the direction of parasite selection in the snail Potamopyrgus antipodarum and its coevolving parasite, Microphallus sp. In the early 2000s, asexuals were more infected than sexuals. A decade later, the asexuals had declined in frequency and were less infected than sexuals. Over time, the mean infection prevalence of asexuals equaled that of sexuals but varied far more. This variation in asexual infection prevalence suggests the potential for parasite-mediated fluctuations in asexual fitness. Accordingly, we detected fitness consequences of the shift in parasite selection: when they were less infected than sexuals, asexuals increased in frequency in the field and in paired mesocosms that isolated the effect of parasites. The match between field and experiment argues that coevolving parasites drive temporal change in the relative fitness and frequency of asexuals, potentially promoting the coexistence of reproductive modes in P. antipodarum.

Bloody-minded parasites and sex: the effects of fluctuating virulence.

Asexual lineages can grow at a faster rate than sexual lineages. Why then is sexual reproduction so widespread? Much empirical evidence supports the Red Queen hypothesis. Under this hypothesis, coevolving parasites favour sexual reproduction by adapting to infect common asexual clones and driving them down in frequency. One limitation, however, seems to challenge the generality of the Red Queen: in theoretical models, parasites must be very virulent to maintain sex. Moreover, experiments show virulence to be unstable, readily shifting in response to environmental conditions. Does variation in virulence further limit the ability of coevolving parasites to maintain sex? To address this question, we simulated temporal variation in virulence and evaluated the outcome of competition between sexual and asexual females. We found that variation in virulence did not limit the ability of coevolving parasites to maintain sex. In fact, relatively high variation in virulence promoted parasite-mediated maintenance of sex. With sufficient variation, sexual females persisted even when mean virulence fell well below the threshold virulence required to maintain sex under constant conditions. We conclude that natural variation in virulence does not limit the relevance of the Red Queen hypothesis for natural populations; on the contrary, it could expand the range of conditions over which coevolving parasites can maintain sex.

50-year anniversary of Lloyd's "mean crowding": Ideas on patchy distributions.

This year marks the 50th anniversary of Monte B. Lloyd's "Mean Crowding" (1967) paper, in which he introduced a metric that accounts for an individual's experience of conspecific density. Mean crowding allows ecologists to measure the degree of spatial aggregation of individuals in a manner relevant to intraspecific competition for resources. We take the concept of mean crowding a step beyond its most common usage and that it has a mathematical relationship to many of the most important concepts in ecology and evolutionary biology. Mean crowding, a first-order approximation of the degree of nonrandomness in a distribution, can function as a powerful heuristic that can unify concepts across disciplines in a more general way that Lloyd originally envisioned.

Habitat Heterogeneity, Host Population Structure, and Parasite Local Adaptation.

Reciprocal-transplant experiments have proven to be a powerful tool for detecting local adaptation (LA). More recently, reciprocal cross-inoculation experiments have been used to evaluate adaptation by parasites to their local host populations. These experiments are conceptually similar to reciprocal-transplant experiments, except that the "environment" (the host population) may have evolved in response to changes in the parasite population. Here, I use analytical tools and computer simulations to determine when parasites would be expected to be more infective to their local host populations than to allopatric host populations. The models assume that parasites have to genetically "match" their hosts in order to infect. I also assumed that different host clones were favored in different populations. When parasite virulence was low, clonal selection outweighed parasite-mediated selection, leading to low host diversity within populations and strong LA by parasites. At intermediate levels of virulence, parasite-mediated selection maintained high levels of host diversity within populations, which reduced or eliminated parasite LA. The loss of parasite LA was not associated with increased infectivity by parasites on allopatric hosts. Instead, the loss of LA was due to a reduction in infectivity of parasites on sympatric hosts. Finally, at high levels of parasite virulence, parasite-mediated selection led to oscillatory host dynamics and weak local adaption by parasites. Across all levels of virulence, the strength of parasite LA closely tracked the degree of host population structure (GST).

Coevolutionary Epidemiology: Disease Spread, Local Adaptation, and Sex.

How does evolution in parasite populations affect the rate of disease spread? In the present study, I derived the mean reproductive rate ([Formula: see text]) for a genetically diverse parasite population that is evolving with a similarly diverse host population. Assuming a matching-alleles model, I found that [Formula: see text] is a positive function of the covariance between the frequencies of "matching" host and parasite genotypes. Computer simulations further showed that evolution in the parasite population tends to increase the covariance, which can lead to epidemiological feedbacks. However, the covariances can also become negative during counteradaptation by the host, leading to oscillatory dynamics in host and parasite fitness. Nonetheless, when parasite-mediated selection is strong, the covariance is positive on average, which facilitates the spread of disease. Positive covariances may also underlie patterns of local adaptation in parasite populations and increase the selective advantage of cross-fertilization in host populations.

Fine-Scale Spatial Covariation between Infection Prevalence and Susceptibility in a Natural Population.

The prevalence of infection varies dramatically on a fine spatial scale. Many evolutionary hypotheses are founded on the assumption that this variation is due to host genetics, such that sites with a high frequency of alleles conferring susceptibility are associated with higher infection prevalence. This assumption is largely untested and may be compromised at finer spatial scales where gene flow between sites is high. We put this assumption to the test in a natural snail-trematode interaction in which host susceptibility is known to have a strong genetic basis. A decade of field sampling revealed substantial spatial variation in infection prevalence between 13 sites around a small lake. Laboratory assays replicated over 3 years demonstrate striking variation in host susceptibility among sites in spite of high levels of gene flow between sites. We find that mean susceptibility can explain more than one-third of the observed variation in mean infection prevalence among sites. We estimate that variation in susceptibility and exposure together can explain the majority of variation in prevalence. Overall, our findings in this natural host-parasite system argue that spatial variation in infection prevalence covaries strongly with variation in the distribution of genetically based susceptibility, even at a fine spatial scale.

Infection dynamics in coexisting sexual and asexual host populations: support for the Red Queen hypothesis.

The persistence of sexual reproduction is a classic problem in evolutionary biology. The problem stems from the fact that, all else equal, asexual lineages should rapidly replace coexisting sexual individuals due to the cost of producing males in sexual populations. One possible countervailing advantage to sexual reproduction is that, on average, outcrossed offspring are more resistant than common clones to coevolving parasites, as predicted under the Red Queen hypothesis. In this study, we evaluated the prevalence of infection by a sterilizing trematode (Microphallus sp.) in a natural population of freshwater snails that was composed of both sexual and asexual individuals (Potamopyrgus antipodarum). More specifically, we compared the frequency of infection in sexual and asexual individuals over a 5-year period at four sites at a natural glacial lake (Lake Alexandrina, South Island, New Zealand). We found that at most sites and over most years, the sexual population was less infected than the coexisting asexual population. Moreover, the frequency of uninfected sexual females was periodically greater than two times the frequency of uninfected asexual females. These results give clear support for a fluctuating parasite-mediated advantage to sexual reproduction in a natural population.

Experimental coevolution: rapid local adaptation by parasites depends on host mating system.

Host-parasite interactions can drive rapid, reciprocal genetic changes (coevolution), provided both hosts and parasites have high heritabilities for resistance/infectivity. Similarly, the host's mating system should also affect the rate of coevolutionary change in host-parasite interactions. Using experimental coevolution, we determined the effect of obligate outcrossing verses partial self-fertilization (mixed mating) on the rate of evolutionary change in a nematode host (Caenorhabditis elegans) and its bacterial parasite (Serratia marcescens). Bacterial populations were derived from a common ancestor. We measured the effects of host mating system on host adaptation to the parasite. We then determined the extent of parasite adaptation to their local host populations. Obligately outcrossing hosts exhibited more rapid adaptation to parasites than did mixed mating hosts. In addition, most of the parasites became adapted to infecting their local hosts, but parasites from obligately outcrossing hosts showed a greater level of local adaptation. These results suggest that host populations evolved along separate trajectories and that outcrossing host populations diverged further than partially selfing populations. Finally, parasites tracking outcrossing host populations diverged further than parasites tracking the partially selfing host populations. These results show that the evolutionary trajectories of both hosts and parasites can be shaped by the host's mating system.

Outcrossing increases resistance against coevolving parasites.

Despite substantial costs, biparental sex is the dominant mode of reproduction across plant and animal taxa. The Red Queen hypothesis (RQH) posits that coevolutionary interactions with parasites can favor biparental sex in hosts, despite the costs. In support of the RQH, previous studies found that coevolutionary interactions with virulent bacterial parasites maintained high outcrossing rates in populations of the androdioecious nematode host Caenorhabditis elegans . Here we test three non-mutually exclusive mechanisms that could explain how coevolving parasites maintain outcrossing rates in C. elegans hosts: 1) short-term parasite exposure induces plastic increases in the hosts' propensity to outcross, 2) hosts evolve increased outcrossing propensity in response to selection imposed by coevolving parasites, and 3) outcrossed offspring incur less parasite-mediated fitness loss than selfed offspring, increasing host male frequencies and opportunities for outcrossing. We find no evidence that parasites cause plastic or evolved changes in host outcrossing propensity. However, parental outcrossing significantly increases survival of host offspring in the F2 generation when exposed to a coevolving parasite. Hence, coevolving parasites maintain outcrossing in host populations by selecting against selfed offspring, rather than by inducing changes in the propensity to outcross.

Outcrossing in Caenorhabditis elegans increases in response to food limitation.

Theory predicts that organisms should diversify their offspring when faced with a stressful environment. This prediction has received empirical support across diverse groups of organisms and stressors. For example, when encountered by Caenorhabditis elegans during early development, food limitation (a common environmental stressor) induces the nematodes to arrest in a developmental stage called dauer and to increase their propensity to outcross when they are subsequently provided with food and enabled to develop to maturity. Here we tested whether food limitation first encountered during late development/early adulthood can also induce increased outcrossing propensity in C. elegans. Previously well-fed C. elegans increased their propensity to outcross when challenged with food limitation during the final larval stage of development and into early adulthood, relative to continuously well-fed (control) nematodes. Our results thus support previous research demonstrating that the stress of food limitation can induce increased outcrossing propensity in C. elegans. Furthermore, our results expand on previous work by showing that food limitation can still increase outcrossing propensity even when it is not encountered until late development, and this can occur independently of the developmental and gene expression changes associated with dauer.

The geographic mosaic of sex and infection in lake populations of a New Zealand snail at multiple spatial scales.

Understanding how sexual and asexual forms of the same species coexist is a challenge for evolutionary biology. The Red Queen hypothesis predicts that sex is favored by parasite-mediated selection against common asexual genotypes, leading to the coexistence of sexual and asexual hosts. In a geographic mosaic, where the risk of infection varies in space, the theory also predicts that sexual reproduction would be positively correlated with disease prevalence. We tested this hypothesis in lake populations of a New Zealand freshwater snail, Potamopyrgus antipodarum, by comparing pairwise difference matrices for infection frequency and male frequency using partial Mantel tests. We conducted the test at three spatial scales: among lakes on the South Island, among depths within an intensively sampled lake (Lake Alexandrina), and within depths at Lake Alexandrina. We found that the difference in infection risk and the difference in the proportion of sexual snails were significantly and positively correlated at all spatial scales. Our results thus suggest that parasite-mediated selection contributes to the long-term coexistence of sexual and asexual individuals in coevolutionary hotspots, and that the "warmth" of hotspots can vary on small spatial scales.

Host Association and Spatial Proximity Shape but Do Not Constrain Population Structure in the Mutualistic Symbiont Xenorhabdus bovienii.

To what extent are generalist species cohesive evolutionary units rather than a compilation of recently diverged lineages? We examine this question in the context of host specificity and geographic structure in the insect pathogen and nematode mutualist Xenorhabdus bovienii. This bacterial species partners with multiple nematode species across two clades in the genus Steinernema. We sequenced the genomes of 42 X. bovienii strains isolated from four different nematode species and three field sites within a 240-km2 region and compared them to globally available reference genomes. We hypothesized that X. bovienii would comprise several host-specific lineages, such that bacterial and nematode phylogenies would be largely congruent. Alternatively, we hypothesized that spatial proximity might be a dominant signal, as increasing geographic distance might lower shared selective pressures and opportunities for gene flow. We found partial support for both hypotheses. Isolates clustered largely by nematode host species but did not strictly match the nematode phylogeny, indicating that shifts in symbiont associations across nematode species and clades have occurred. Furthermore, both genetic similarity and gene flow decreased with geographic distance across nematode species, suggesting differentiation and constraints on gene flow across both factors, although no absolute barriers to gene flow were observed across the regional isolates. Several genes associated with biotic interactions were found to be undergoing selective sweeps within this regional population. The interactions included several insect toxins and genes implicated in microbial competition. Thus, gene flow maintains cohesiveness across host associations in this symbiont and may facilitate adaptive responses to a multipartite selective environment. IMPORTANCE Microbial populations and species are notoriously hard to delineate. We used a population genomics approach to examine the population structure and the spatial scale of gene flow in Xenorhabdus bovienii, an intriguing species that is both a specialized mutualistic symbiont of nematodes and a broadly virulent insect pathogen. We found a strong signature of nematode host association, as well as evidence for gene flow connecting isolates associated with different nematode host species and collected from distinct study sites. Furthermore, we saw signatures of selective sweeps for genes involved with nematode host associations, insect pathogenicity, and microbial competition. Thus, X. bovienii exemplifies the growing consensus that recombination not only maintains cohesion but can also allow the spread of niche-beneficial alleles.

Host-parasite coevolution: Partitioning the effects of natural selection and environmental change using coupled Price equations.

George Price showed how the effects of natural selection and environmental change could be mathematically partitioned. This partitioning may be especially useful for understanding host-parasite coevolution, where each species represents the environment for the other species. Here, we use coupled Price equations to study this kind of antagonistic coevolution. We made the common assumption that parasites must genetically match their host's genotype to avoid detection by the host's self/nonself recognition system, but we allowed for the possibility that non-matching parasites have some fitness. Our results show how natural selection on one species results in environmental change for the other species. Numerical iterations of the model show that these environmental changes can periodically exceed the changes in mean fitness due to natural selection, as suggested by R.A. Fisher. Taken together, the results give an algebraic dissection of the eco-evolutionary feedbacks created during host-parasite coevolution.

Trans-specific polymorphism and the convergent evolution of supertypes in major histocompatibility complex class II genes in darters (Etheostoma).

Major Histocompatibility Complex (MHC) genes are one of the most polymorphic gene groups known in vertebrates. MHC genes also exhibit allelic variants that are shared among taxa, referred to as trans-specific polymorphism (TSP). The role that selection plays in maintaining such high diversity within species, as well as TSP, is an ongoing discussion in biology. In this study, we used deep-sequencing techniques to characterize MHC class IIb gene diversity in three sympatric species of darters. We found at least 5 copies of the MHC gene in darters, with 126 genetic variants encoding 122 unique amino acid sequences. We identified four supertypes based on the binding properties of proteins encoded by the sequences. Although each species had a unique pool of variants, many variants were shared between species pairs and across all three species. Phylogenetic analysis showed that the variants did not group together monophyletically based on species identity or on supertype. An expanded phylogenetic analysis showed that some darter alleles grouped together with alleles from other percid fishes. Our findings show that TSP occurs in darters, which suggests that balancing selection is acting at the genotype level. Supertypes, however, are most likely evolving convergently, as evidenced by the fact that alleles do not form monophyletic groups based on supertype. Our research demonstrates that selection may be acting differently on MHC genes at the genotype and supertype levels, selecting for the maintenance of high genotypic diversity while driving the convergent evolution of similar MHC phenotypes across different species.