The Emergence of Non-Linear Evolutionary Trade-offs and the Maintenance of Genetic Polymorphisms.

Evolutionary models of quantitative traits often assume trade-offs between beneficial and detrimental traits, requiring modelers to specify a function linking costs to benefits. The choice of trade-off function is often consequential; functions that assume diminishing returns (accelerating costs) typically lead to single equilibrium genotypes, while decelerating costs often lead to evolutionary branching. Despite their importance, we still lack a strong theoretical foundation to base the choice of trade-off function. To address this gap, we explore how trade-off functions can emerge from the genetic architecture of a quantitative trait. We developed a multi-locus model of disease resistance, assuming each locus had random antagonistic pleiotropic effects on resistance and fecundity. We used this model to generate genotype landscapes and explored how additive versus epistatic genetic architectures influenced the shape of the trade-off function. Regardless of epistasis, our model consistently led to accelerating costs. We then used our genotype landscapes to build an evolutionary model of disease resistance. Unlike other models with accelerating costs, our approach often led to genetic polymorphisms at equilibrium. Our results suggest that accelerating costs are a strong null model for evolutionary trade-offs and that the eco-evolutionary conditions required for polymorphism may be more nuanced than previously believed.

High temperatures reduce growth, infection, and transmission of a naturally occurring fungal plant pathogen.

Climate change is rapidly altering the distribution of suitable habitats for many species as well as their pathogenic microbes. For many pathogens, including vector-borne diseases of humans and agricultural pathogens, climate change is expected to increase transmission and lead to pathogen range expansions. However, if pathogens have a lower heat tolerance than their host, increased warming could generate so-called thermal refugia for hosts. Predicting the outcomes of warming on disease transmission requires detailed knowledge of the thermal tolerances of both the host and the pathogen. Such thermal tolerance studies are generally lacking for fungal pathogens of wild plant populations, despite the fact that plants form the base of all terrestrial communities. Here, we quantified three aspects of the thermal tolerance (growth, infection, and propagule production) of the naturally occurring fungal pathogen Microbotryum lychnidis-dioicae, which causes a sterilizing anther-smut disease on the herbaceous plant Silene latifolia. We also quantified two aspects of host thermal tolerance: seedling survival and flowering rate. We found that temperatures >30°C reduced the ability of anther-smut spores to germinate, grow, and conjugate in vitro. In addition, we found that high temperatures (30°C) during or shortly after the time of inoculation strongly reduced the likelihood of infection in seedlings. Finally, we found that high summer temperatures in the field temporarily cured infected plants, likely reducing transmission. Notably, high temperatures did not reduce survival or flowering of the host plants. Taken together, our results show that the fungus is considerably more sensitive to high temperatures than its host plant. A warming climate could therefore result in reduced disease spread or even local pathogen extirpation, leading to thermal refugia for the host.

Host-pathogen coevolution promotes the evolution of general, broad-spectrum resistance and reduces foreign pathogen spillover risk.

Genetic variation for disease resistance within host populations can strongly impact the spread of endemic pathogens. In plants, recent work has shown that within-population variation in resistance can also affect the transmission of foreign spillover pathogens if that resistance is general. However, most hosts also possess specific resistance mechanisms that provide strong defenses against coevolved endemic pathogens. Here we use a modeling approach to ask how antagonistic coevolution between hosts and their endemic pathogen at the specific resistance locus can affect the frequency of general resistance, and therefore a host's vulnerability to foreign pathogens. We develop a two-locus model with variable recombination that incorporates both general resistance (effective against all pathogens) and specific resistance (effective against endemic pathogens only). With coevolution, when pathogens can evolve to evade specific resistance, we find that the regions where general resistance can evolve are greatly expanded, decreasing the risk of foreign pathogen invasion. Furthermore, coevolution greatly expands the conditions that maintain polymorphisms at both resistance loci, thereby driving greater genetic diversity within host populations. This genetic diversity often leads to positive correlations between host resistance to foreign and endemic pathogens, similar to those observed in natural populations. However, if resistance loci become linked, the resistance correlations can shift to negative. If we include a third linkage-modifying locus in our model, we find that selection often favors complete linkage. Our model demonstrates how coevolutionary dynamics with an endemic pathogen can mold the resistance structure of host populations in ways that affect its susceptibility to foreign pathogen spillovers, and that the nature of these outcomes depends on resistance costs, as well as the degree of linkage between resistance genes.

Quantitative disease resistance in wild Silene vulgaris to its endemic pathogen Microbotryum silenes-inflatae.

The evolution of disease resistances is an expected feature of plant-pathogen systems, but whether the genetics of this trait most often produces qualitative or quantitative phenotypic variation is a significant gap in our understanding of natural populations. These two forms of resistance variation are often associated with differences in number of underlying loci, the specificities of host-pathogen coevolution, as well as contrasting mechanisms of preventing or slowing the infection process. Anther-smut disease is a commonly studied model for disease of wild species, where infection has severe fitness impacts, and prior studies have suggested resistance variation in several host species. However, because the outcome of exposing the individual host to this pathogen is binary (healthy or diseased), resistance has been previously measured at the family level, as the proportion of siblings that become diseased. This leaves uncertain whether among-family variation reflects contrasting ratios of segregating discrete phenotypes or continuous trait variation among individuals. In the host Silene vulgaris, plants were replicated by vegetative propagation in order to quantify the infection rates of the individual genotype with the endemic anther-smut pathogen, Microbotryum silenes-inflatae. The variance among field-collected families for disease resistance was significant, while there was unimodal continuous variation in resistance among genotypes. Using crosses between genotypes within ranked resistance quartiles, the offspring infection rate was predicted by the parental resistance values. While the potential remains in this system for resistance genes having major effects, as there were suggestions of such qualitative resistance in a prior study, here the quantitative disease resistance to the endemic anther-smut pathogen is indicated for S. vulgaris. The variation in natural populations and strong heritability of the trait, combined with severe fitness consequences of anther-smut disease, suggests that resistance in these host populations is highly capable of responding to disease-induced selection.

Host-pathogen coevolution promotes the evolution of general, broad-spectrum resistance and reduces foreign pathogen spillover risk.

Genetic variation for disease resistance within host populations can strongly impact the spread of endemic pathogens. In plants, recent work has shown that within-population variation in resistance can also affect the transmission of foreign spillover pathogens if that resistance is general. However, most hosts also possess specific resistance mechanisms that provide strong defenses against coevolved endemic pathogens. Here we use a modeling approach to ask how antagonistic coevolution between hosts and their endemic pathogen at the specific resistance locus can affect the frequency of general resistance, and therefore a host's vulnerability to foreign pathogens. We develop a two-locus model with variable recombination that incorporates both general (resistance to all pathogens) and specific (resistance to endemic pathogens only). We find that introducing coevolution into our model greatly expands the regions where general resistance can evolve, decreasing the risk of foreign pathogen invasion. Furthermore, coevolution greatly expands which conditions maintain polymorphisms at both resistance loci, thereby driving greater genetic diversity within host populations. This genetic diversity often leads to positive correlations between host resistance to foreign and endemic pathogens, similar to those observed in natural populations. However, if resistance loci become linked, the resistance correlations can shift to negative. If we include a third, linkage modifying locus into our model, we find that selection often favors complete linkage. Our model demonstrates how coevolutionary dynamics with an endemic pathogen can mold the resistance structure of host populations in ways that affect its susceptibility to foreign pathogen spillovers, and that the nature of these outcomes depends on resistance costs, as well as the degree of linkage between resistance genes.

Specific resistance prevents the evolution of general resistance and facilitates disease emergence.

Host-shifts, where pathogens jump from an ancestral host to a novel host, can be facilitated or impeded by standing variation in disease resistance, but only if resistance provides broad-spectrum general resistance against multiple pathogen species. Host resistance comes in many forms and includes both general resistance, as well as specific resistance, which may only be effective against a single pathogen species or even genotype. However, most evolutionary models consider only one of these forms of resistance, and we have less understanding of how these two forms of resistance evolve in tandem. Here, we develop a model that allows for the joint evolution of specific and general resistance and asks if the evolution of specific resistance drives a decrease in the evolution of general resistance. We also explore how these evolutionary outcomes affect the risk of foreign pathogen invasion and persistence. We show that in the presence of a single endemic pathogen, the two forms of resistance are strongly exclusionary. Critically, we find that specific resistance polymorphisms can prevent the evolution of general resistance, facilitating the invasion of foreign pathogens. We also show that specific resistance polymorphisms are a necessary condition for the successful establishment of foreign pathogens following invasion, as they prevent the exclusion of the foreign pathogen by the more transmissible endemic pathogen. Our results demonstrate the importance of considering the joint evolution of multiple forms of resistance when evaluating a population's susceptibility to foreign pathogens.

Extinct in the wild: The precarious state of Earth's most threatened group of species.

Extinct in the Wild (EW) species are placed at the highest risk of extinction under the International Union for Conservation of Nature Red List, but the extent and variation in this risk have never been evaluated. Harnessing global databases of ex situ animal and plant holdings, we report on the perilous state of EW species. Most EW animal species-already compromised by their small number of founders-are maintained at population sizes far below the thresholds necessary to ensure demographic security. Most EW plant species depend on live propagation by a small number of botanic gardens, with a minority secured at seed bank institutions. Both extinctions and recoveries are possible fates for EW species. We urgently call for international effort to enable the latter.

The evolution of age-specific resistance to infectious disease.

Innate, infection-preventing resistance often varies between host life stages. Juveniles are more resistant than adults in some species, whereas the opposite pattern is true in others. This variation cannot always be explained by prior exposure or physiological constraints and so it has been hypothesized that trade-offs with other life-history traits may be involved. However, little is known about how trade-offs between various life-history traits and resistance at different life stages affect the evolution of age-specific resistance. Here, we use a mathematical model to explore how trade-offs with natural mortality, reproduction and maturation combine to affect the evolution of resistance at different life stages. Our results show that certain combinations of trade-offs have substantial effects on whether adults or juveniles are more resistant, with trade-offs between juvenile resistance and adult reproduction inherently more costly than trade-offs involving maturation or mortality (all else being equal), resulting in consistent evolution of lower resistance at the juvenile stage even when infection causes a lifelong fecundity reduction. Our model demonstrates how the differences between patterns of age-structured resistance seen in nature may be explained by variation in the trade-offs involved and our results suggest conditions under which trade-offs tend to select for lower resistance in juveniles than adults.

Host density shapes the relative contribution of vector-based and aerial transmission of a pathogenic fungus.

Pathogen transmission mode is a key determinant of epidemiological outcomes. Theory shows that host density can influence the spread of pathogens differentially depending on their mode of transmission. Host density could therefore play an important role in determining the pathogen transmission mode. We tested theoretical expectations using floral arrays of the alpine carnation Dianthus pavonius in field experiments of spore dispersal of the anther-smut fungus, Microbotryum, by vector (pollinator)-based floral transmission and passive aerial transmission at a range of host densities. Pollinators deposited fewer spores per plant at high host density than at lower density (ranging from a 0.2-2 m spacing between plants), and vector-based spore deposition at higher densities declined more steeply with distance from diseased plant sources. In contrast, while aerial spore deposition declined with distance from the diseased source, the steepness of this decline was independent of host density. Our study indicates that the amount and distance of vector-based transmission are likely to be a nonmonotonic function of host density as a result of vector behavior, which is not readily encapsulated by fixed dispersal functions. We conclude that the spatial spread of pathogens by vectors is likely to be greater at lower and intermediate densities, whereas the spatial spread of aerially transmitted pathogens would be greater at high densities. These contrasting patterns could lead to differential importance of each transmission mode in terms of its contribution to subsequent infections across host densities.

Multimodal pathogen transmission as a limiting factor in host distribution.

Theoretical models suggest that infectious diseases could play a substantial role in determining the spatial extent of host species, but few studies have collected the empirical data required to test this hypothesis. Pathogens that sterilize their hosts or spread through frequency-dependent transmission could have especially strong effects on the limits of species' distributions because diseased hosts that are sterilized but not killed may continue to produce infectious stages and frequency-dependent transmission mechanisms are effective even at very low population densities. We collected spatial pathogen prevalence data and population abundance data for alpine carnations infected by the sterilizing pathogen Microbotryum dianthorum, a parasite that is spread through both frequency-dependent (vector-borne) and density-dependent (aerial spore transmission) mechanisms. Our 13-year study reveals rapid declines in population abundance without a compensatory decrease in pathogen prevalence. We apply a stochastic, spatial model of parasite spread that accommodates spatial habitat heterogeneity to investigate how the population dynamics depend on multimodal (frequency-dependent and density-dependent) transmission. We found that the observed rate of population decline could plausibly be explained by multimodal transmission, but is unlikely to be explained by either frequency-dependent or density-dependent mechanisms alone. Multimodal pathogen transmission rates high enough to explain the observed decline predicted that eventual local extinction of the host species is highly likely. Our results add to a growing body of literature showing how multimodal transmission can constrain species distributions in nature.

Can disease resistance evolve independently at different ages? Genetic variation in age-dependent resistance to disease in three wild plant species.

Juveniles are typically less resistant (more susceptible) to infectious disease than adults, and this difference in susceptibility can help fuel the spread of pathogens in age-structured populations. However, evolutionary explanations for this variation in resistance across age remain to be tested.One hypothesis is that natural selection has optimized resistance to peak at ages where disease exposure is greatest. A central assumption of this hypothesis is that hosts have the capacity to evolve resistance independently at different ages. This would mean that host populations have (a) standing genetic variation in resistance at both juvenile and adult stages, and (b) that this variation is not strongly correlated between age classes so that selection acting at one age does not produce a correlated response at the other age.Here we evaluated the capacity of three wild plant species (Silene latifolia, S. vulgaris and Dianthus pavonius) to evolve resistance to their anther-smut pathogens (Microbotryum fungi), independently at different ages. The pathogen is pollinator transmitted, and thus exposure risk is considered to be highest at the adult flowering stage.Within each species we grew families to different ages, inoculated individuals with anther smut, and evaluated the effects of age, family and their interaction on infection.In two of the plant species, S. latifolia and D. pavonius, resistance to smut at the juvenile stage was not correlated with resistance to smut at the adult stage. In all three species, we show there are significant age × family interaction effects, indicating that age specificity of resistance varies among the plant families. Synthesis. These results indicate that different mechanisms likely underlie resistance at juvenile and adult stages and support the hypothesis that resistance can evolve independently in response to differing selection pressures as hosts age. Taken together our results provide new insight into the structure of genetic variation in age-dependent resistance in three well-studied wild host-pathogen systems.

Assessing ex situ genetic and ecogeographic conservation in a threatened but widespread oak after range-wide collecting effort.

Although the genetic diversity and structure of in situ populations has been investigated in thousands of studies, the genetic composition of ex situ plant populations has rarely been studied. A better understanding of how much genetic diversity is conserved ex situ, how it is distributed among locations (e.g., botanic gardens), and what minimum sample sizes are needed is necessary to improve conservation outcomes. Here we address these issues in a threatened desert oak species, Quercus havardii Rydb. We assess the genetic, geographic, and ecological representation of 290 plants from eight ex situ locations, relative to 667 wild individuals from 35 in situ locations. We also leverage a recent dataset of >3000 samples from 11 other threatened plants to directly compare the degree of genetic conservation for species that differ in geographic range size. We found that a majority of Q. havardii genetic diversity is conserved; one of its geographic regions is significantly better conserved than the other; genetic diversity conservation of this widespread species is lower than documented for the 11 rarer taxa; genetic diversity within each garden is strongly correlated to the number of plants and number of source populations; and measures of geographic and ecological conservation (i.e., percent area and percent of ecoregions represented) were typically lower than the direct assessment of genetic diversity (i.e., percent alleles). This information will inform future seed sampling expeditions to ensure that the intraspecific diversity of threatened plants can be effectively conserved.

The Tip of the Iceberg: Cryopreservation Needs for Meeting the Challenge of Exceptional Plant Conservation.

Cryopreservation is increasingly important as a conservation tool, particularly for threatened exceptional species. The goal of this study was to investigate the current knowledge of plant cryopreservation through a search of the literature in Web of Science and align that with the 775 species currently identified on the Working List of Exceptional Plants. While there is a good foundation in plant cryopreservation research, particularly with economically important species, there are significant gaps in research on families that contain the largest numbers of currently known exceptional species, including the Dipterocarpaceae, Rhizophoraceae, and Pittosporaceae. Even families well represented in both in the literature and on the List of Exceptional Plants had much less overlap at the level of genus. Tropical trees, a significant portion of exceptional species, were not as well represented in the literature as herbaceous species. Over 70% of all articles dealt with in vitro cryopreservation, with much less emphasis on other methods (seed, embryo, dormant bud, and pollen) that will be more cost-effective for species where they can be applied. While the research on plant cryopreservation to date provides a strong foundation and is being utilized effectively for conserving the diversity of a number of economically important species, this study revealed significant gaps that can help prioritize future research to more effectively conserve the diversity of threatened exceptional species.

Resistance Correlations Influence Infection by Foreign Pathogens.

AbstractReciprocal selection promotes the specificity of host-pathogen associations and resistance polymorphisms in response to disease. However, plants and animals also vary in response to pathogen species not previously encountered in nature, with potential effects on new disease emergence. Using anther smut disease, we show that resistance (measured as infection rates) to foreign pathogens can be correlated with standing variation in resistance to an endemic pathogen. In Silene vulgaris, genetic variation in resistance to its endemic anther smut pathogen correlated positively with resistance variation to an anther smut pathogen from another host, but the relationship was negative between anther smut and a necrotrophic pathogen. We present models describing the genetic basis for assessing resistance relationships between endemic and foreign pathogens and for quantifying infection probabilities on foreign pathogen introduction. We show that even when the foreign pathogen has a lower average infection ability than the endemic pathogen, infection outcomes are determined by the sign and strength of the regression of the host's genetic variation in infection rates by a foreign pathogen on variation in infection rates by an endemic pathogen as well as by resistance allele frequencies. Given that preinvasion equilibria of resistance are determined by factors including resistance costs, we show that protection against foreign pathogens afforded by positively correlated resistances can be lessened or even result in elevated infection risk at the population level, depending on local dynamics. Therefore, a pathogen's emergence potential could be influenced not only by its average infection rate but also by resistance variation resulting from prior selection imposed by endemic diseases.

From generalist to specialists: Variation in the host range and performance of anther-smut pathogens on Dianthus.

Determining the processes that drive the evolution of pathogen host range can inform our understanding of disease dynamics and the potential for host shifts. In natural populations, patterns of host range could be driven by genetically based differences in pathogen infectivity or ecological differences in host availability. In northwestern Italy, four reproductively isolated lineages of the fungal plant-pathogen Microbotryum have been shown to co-occur on several species in the genus Dianthus. We carried out cross-inoculation experiments to determine whether patterns of realized host range in these four lineages were driven by differences in infectivity and to test whether there was evidence of a trade-off between host range and within-host reproduction. We found strong concordance between field patterns of host range and pathogen infectivity on different Dianthus species using experimental inoculation, indicating that infection ability is a major driving force of host range. However, we found no evidence of a trade-off between the ability to infect a wider range of host species and spore production on a shared host.

[Changes in sexual interests and experiences during the COVID-19 pandemic - A qualitative content analysis]. / Veränderungen sexueller Interessen und Erfahrungen während der COVID-19-Pandemie - Eine qualitative Inhaltsanalyse.

BACKGROUND: Contact restrictions imposed to slow the spread of the coronavirus disease 2019 (COVID-19) pandemic affect people's social lives at various levels as well as their mental and sexual health. OBJECTIVE: The present study aimed to assess changes in sexual interests and experiences of residents in Germany during the first wave of social contact restrictions in early 2020. MATERIAL AND METHODS: In an anonymous online survey, answers to an open question regarding changes in sexuality due to contact restrictions were collected and subjected to Mayring's qualitative content analysis. RESULTS: The changes reported by 248 participants could be subsumed under the categories "sexual desire and arousability", "relationship", "sexual activities", "sexual interests and attitudes", "virtual sexual activities", "dating" and "sexual problems and dysfunctions". CONCLUSION: Social distancing rules and restrictions brought about by the COVID-19 pandemic have led to changes in sexuality and relationships, which should be considered in sexual and couple therapy settings.

Vector preference and heterogeneity in host sex ratio can affect pathogen spread in natural plant populations.

Vector-borne diseases threaten human and agricultural health and are a critical component of the ecology of plants and animals. While previous studies have shown that pathogen spread can be affected by vector preferences for host infection status, less attention has been paid to vector preference for host sex, despite abundant evidence of sex-specific variation in disease burden. We investigated vector preference for host infection status and sex in the sterilizing "anther-smut" pathogen (Microbotryum) of the alpine carnation, Dianthus pavonius. The pathogen is transferred among hosts by pollinators that visit infected flowers and become contaminated with spores produced by infected anthers. The host plant has a mixed breeding system with hermaphrodites and females. In experimental floral arrays, pollinators strongly preferred healthy hermaphrodites over both females and diseased plants, consistently across different guilds of pollinators and over multiple years. Using an agent-based model, we showed that pollinator preferences for sex can affect pathogen spread in populations with variable sex ratios, even if there is no preference for infection status. Our results demonstrate that vector preferences for host traits other than infection status can play a critical role in pathogen transmission dynamics when there is heterogeneity for those traits in the host population.

The role of infectious disease in the evolution of females: Evidence from anther-smut disease on a gynodioecious alpine carnation.

In flowering plants, the evolution of females is widely hypothesized to be the first step in the evolutionary pathway to separate male and female sexes, or dioecy. Natural enemies have the potential to drive this evolution if they preferentially attack hermaphrodites over females. We studied sex-based differences in exposure to anther-smut (Microbotryum), a sterilizing pollinator-transmitted disease, in Dianthus pavonius, a gynodioecious perennial herb. We found that within a heavily diseased population, females consistently had lower levels of Microbotryum spore deposition relative to hermaphrodites and that this difference was driven by rapid floral closing in females following successful pollination. We further show that this protective closing behavior is frequency dependent; females close faster when they are rare. These results indicate that anther-smut disease is an important source of selection for females, especially since we found in a common garden experiment no evidence that females have any inherent fecundity advantages over hermaphrodites. Finally, we show that among populations, those where anther-smut is present have a significantly higher frequency of females than those where the disease is absent. Taken together our results indicate that anther-smut disease is likely an important biotic factor driving the evolution and maintenance of females in this gynodioecious species.

Mitigation of Secondary Organic Aerosol Formation from Log Wood Burning Emissions by Catalytic Removal of Aromatic Hydrocarbons.

Log wood burning is a significant source of volatile organic compounds including aromatic hydrocarbons (ArHC). ArHC are harmful, are reactive in the ambient atmosphere, and are important secondary organic aerosol (SOA) precursors. Consequently, SOA represents a major fraction of the sub-micron organic aerosol pollution from log wood burning. ArHC reduction is thus critical in the mitigation of adverse health and environmental effects of log wood burning. In this study, two Pt-based catalytic converters were prepared and tested for the mitigation of real-world log wood burning emissions, including ArHC and SOA formation, as well as toxic carbon monoxide and methane, a greenhouse gas. Substantial removal of mono- and polycyclic ArHC and phenolic compounds was achieved with both catalysts operated at realistic chimney temperatures (50% conversion was achieved at 200 and 300 °C for non-methane hydrocarbons in our experiments for Pt/Al2O3 and Pt/CeO2-Al2O3, respectively). The catalytically cleaned emissions exhibited a substantially reduced SOA formation already at temperatures as low as 185-310 °C. This reduces the sub-micron PM burden of log wood burning significantly. Thus, catalytic converters can effectively reduce primary and secondary log wood burning pollutants and, thereby, their adverse health impacts and environmental effects.

Effect of the anther-smut fungus Microbotryum on the juvenile growth of its host Silene latifolia.

PREMISE OF THE STUDY: Plant pathogens that form persistent systemic infections within plants have the potential to affect multiple plant life history traits, yet we tend to focus only on visible symptoms. Anther smut of Silene latifolia caused by the fungus Microbotryum lychnidis-dioicae induces the anthers of its host to support fungal spore production instead of pollen, and the pathogen is primarily transmitted among flowering plants by pollinators. Nevertheless, most of its life cycle is spent in the asymptomatic vegetative phase, and spores falling on seedlings or nonflowering plants can also infect the host. The purpose of this study was to ask whether the fungus also had an effect on its host plant in the juvenile vegetative phase before flowering as this is important for the disease dynamics in species where infection of seedlings is commonplace. METHODS: Leaf length and leaf number of inoculated and uninoculated juvenile plants were compared in greenhouse experiments, and in one experiment, disease status of the plants at flowering was determined. KEY RESULTS: Inoculated plants had shorter but more leaves, and reduced root mass at the early juvenile (preflowering) stage. Some of these effects were detectable in plants that were inoculated but showed no disease symptoms at flowering. CONCLUSIONS: These results show that pathogenic fungi can have endophyte-like effects even in the total absence of their typical and more charismatic symptoms, and conversely that the assessment of endophyte effects on the fitness of their hosts should include all stages of the host life cycle.