Elevated atmospheric concentrations of carbon dioxide reduce monarch tolerance and increase parasite virulence by altering the medicinal properties of milkweeds

Hosts combat their parasites using mechanisms of resistance and tolerance, which together determine parasite virulence. Environmental factors, including diet, mediate the impact of parasites on hosts, with diet providing nutritional and medicinal properties. Here, we present the first evidence that ongoing environmental change decreases host tolerance and increases parasite virulence through a loss of dietary medicinal quality. Monarch butterflies use dietary toxins (cardenolides) to reduce the deleterious impacts of a protozoan parasite. We fed monarch larvae foliage from four milkweed species grown under either elevated or ambient CO₂, and measured changes in resistance, tolerance, and virulence. The most high‐cardenolide milkweed species lost its medicinal properties under elevated CO₂; monarch tolerance to infection decreased, and parasite virulence increased. Declines in medicinal quality were associated with declines in foliar concentrations of lipophilic cardenolides. Our results emphasize that global environmental change may influence parasite–host interactions through changes in the medicinal properties of plants.     

Genetic variation in resistance, but not tolerance, to a protozoan parasite in the monarch butterfly

Natural selection should strongly favour hosts that can protect themselves against parasites. Most studies on animals so far have focused on resistance, a series of mechanisms through which hosts prevent infection, reduce parasite growth or clear infection. However, animals may instead evolve tolerance, a defence mechanism by which hosts do not reduce parasite infection or growth, but instead alleviate the negative fitness consequences of such infection and growth. Here, we studied genetic variation in resistance and tolerance in the monarch butterfly (Danaus plexippus) to its naturally occurring protozoan parasite, Ophryocystis elektroscirrha. We exposed 560 monarch larvae of 19 different family lines to one of five different parasite inoculation doses (0, 1, 5, 10 and 100 infective spores) to create a range of parasite loads in infected butterflies. We then used two proxies of host fitness (adult lifespan and body mass) to quantify: (i) qualitative resistance (the ability to prevent infection; also known as avoidance or anti-infection resistance); (ii) quantitative resistance (the ability to limit parasite growth upon infection; also known as control or anti-growth resistance); and (iii) tolerance (the ability to maintain fitness with increasing parasite infection intensity). We found significant differences among host families in qualitative and quantitative resistance, indicating genetic variation in resistance. However, we found no genetic variation in tolerance. This may indicate that all butterflies in our studied population have evolved maximum tolerance, as predicted by some theoretical models.     

Behavioural resistance against a protozoan parasite in the monarch butterfly

1. As parasites can dramatically reduce the fitness of their hosts, there should be strong selection for hosts to evolve and maintain defence mechanisms against their parasites. One way in which hosts may protect themselves against parasitism is through altered behaviours, but such defences have been much less studied than other forms of parasite resistance. 2. We studied whether monarch butterflies (Danaus plexippus L.) use altered behaviours to protect themselves and their offspring against the protozoan parasite Ophryocystis elektroscirrha (McLaughlin & Myers (1970), Journal of Protozoology, 17, p. 300). In particular, we studied whether (i) monarch larvae can avoid contact with infectious parasite spores; (ii) infected larvae preferentially consume therapeutic food plants when given a choice or increase the intake of such plants in the absence of choice; and (iii) infected female butterflies preferentially lay their eggs on medicinal plants that make their offspring less sick. 3. We found that monarch larvae were unable to avoid infectious parasite spores. Larvae were also not able to preferentially feed on therapeutic food plants or increase the ingestion of such plants. However, infected female butterflies preferentially laid their eggs on food plants that reduce parasite growth in their offspring. 4. Our results suggest that animals may use altered behaviours as a protection against parasites and that such behaviours may be limited to a single stage in the host-parasite life cycle. Our results also suggest that animals may use altered behaviours to protect their offspring instead of themselves. Thus, our study indicates that an inclusive fitness approach should be adopted to study behavioural defences against parasites.     

Aphids indirectly increase virulence and transmission potential of a monarch butterfly parasite by reducing defensive chemistry of a shared food plant

Parasites and hosts live in communities consisting of many interacting species, but few studies have examined how communities affect parasite virulence and transmission. We studied a food web consisting of two species of milkweed, two milkweed herbivores (monarch butterfly and oleander aphid) and a monarch butterfly-specific parasite. We found that the presence of aphids increased the virulence and transmission potential of the monarch butterfly's parasite on one milkweed species. These increases were associated with aphid-induced decreases in the defensive chemicals of milkweed plants. Our experiment suggests that aphids can indirectly increase the virulence and transmission potential of monarch butterfly parasites, probably by altering the chemical composition of a shared food plant. These results indicate that species that are far removed from host-parasite interactions can alter such interactions through cascading indirect effects in the food web. As such, indirect effects within ecological communities may drive the dynamics and evolution of parasites.     

Sex‐dependent infection causes nonadditive effects on kissing bug fecundity

The influence of parasites on host reproduction has been widely studied in natural and experimental conditions. Most studies, however, have evaluated the parasite impact on female hosts only, neglecting the contribution of males for host reproduction. This omission is unfortunate as sex‐dependent infection may have important implications for host–parasite associations. Here, we evaluate for the first time the independent and nonindependent effects of gender infection on host reproductive success using the kissing bug Mepraia spinolai and the protozoan Trypanosoma cruzi as model system. We set up four crossing treatments including the following: (1) both genders infected, (2) both genders uninfected, (3) males infected—females uninfected, and (4) males uninfected—females infected, using fecundity measures as response variables. Interactive effects of infection between sexes were prevalent. Uninfected females produced more and heavier eggs when crossed with uninfected than infected males. Uninfected males, in turn, sired more eggs and nymphs when crossed with uninfected than infected females. Unexpectedly, infected males sired more nymphs when crossed with infected than uninfected females. These results can be explained by the effect of parasitism on host body size. As infection reduced size in both genders, infection on one sex only creates body size mismatches and mating constraints that are not present in pairs with the same infection status. Our results indicate the fitness impact of parasitism was contingent on the infection status of genders and mediated by body size. As the fecundity impact of parasitism cannot be estimated independently for each gender, inferences based only on female host infection run the risk of providing biased estimates of parasite‐mediated impact on host reproduction.     

Differential Tolerance to Direct and Indirect Density-Dependent Costs of Viral Infection in Arabidopsis thaliana

Population density and costs of parasite infection may condition the capacity of organisms to grow, survive and reproduce, i.e. their competitive ability. In host–parasite systems there are different competitive interactions: among uninfected hosts, among infected hosts, and between uninfected and infected hosts. Consequently, parasite infection results in a direct cost, due to parasitism itself, and in an indirect cost, due to modification of the competitive ability of the infected host. Theory predicts that host fitness reduction will be higher under the combined effects of costs of parasitism and competition than under each factor separately. However, experimental support for this prediction is scarce, and derives mostly from animal–parasite systems. We have analysed the interaction between parasite infection and plant density using the plant-parasite system of Arabidopsis thaliana and the generalist virus Cucumber mosaic virus (CMV). Plants of three wild genotypes grown at different densities were infected by CMV at various prevalences, and the effects of infection on plant growth and reproduction were quantified. Results demonstrate that the combined effects of host density and parasite infection may result either in a reduction or in an increase of the competitive ability of the host. The two genotypes investing a higher proportion of resources to reproduction showed tolerance to the direct cost of infection, while the genotype investing a higher proportion of resources to growth showed tolerance to the indirect cost of infection. Our findings show that the outcome of the interaction between host density and parasitism depends on the host genotype, which determines the plasticity of life-history traits and consequently, the host capacity to develop different tolerance mechanisms to the direct or indirect costs of parasitism. These results indicate the high relevance of host density and parasitism in determining the competitive ability of a plant, and stress the need to simultaneously consider both factors to understand the selective pressures that drive host–parasite co-evolution.     

STEPWISE EVOLUTION OF RESISTANCE TO TOXIC CARDENOLIDES VIA GENETIC SUBSTITUTIONS IN THE NA+/K+‐ATPASE OF MILKWEED BUTTERFLIES (LEPIDOPTERA: DANAINI)

Despite the monarch butterfly (Danaus plexippus) being famous for its adaptations to the defensive traits of its milkweed host plants, little is known about the macroevolution of these traits. Unlike most other animal species, monarchs are largely insensitive to cardenolides, because their target site, the sodium pump (Na⁺/K⁺‐ATPase), has evolved amino acid substitutions that reduce cardenolide binding (so‐called target site insensitivity, TSI). Because many, but not all, species of milkweed butterflies (Danaini) are associated with cardenolide‐containing host plants, we analyzed 16 species, representing all phylogenetic lineages of milkweed butterflies, for the occurrence of TSI by sequence analyses of the Na⁺/K⁺‐ATPase gene and by enzymatic assays with extracted Na⁺/K⁺‐ATPase. Here we report that sensitivity to cardenolides was reduced in a stepwise manner during the macroevolution of milkweed butterflies. Strikingly, not all Danaini typically consuming cardenolides showed TSI, but rather TSI was more strongly associated with sequestration of toxic cardenolides. Thus, the interplay between bottom‐up selection by plant compounds and top‐down selection by natural enemies can explain the evolutionary sequence of adaptations to these toxins.     

Infection risk dictates immunological divergence among populations in a Mediterranean lizard

The ability of vertebrates to evolve different defence strategies in response to varying parasitism regimes remains poorly understood. Hosts may adopt two different strategies to defend themselves against parasites: tolerance (hosts alleviate the negative fitness consequences of parasite infection) and resistance (hosts strengthen their immune response as parasite burden increases). Both strategies are effective, but fitness has been reported to decline faster in less‐tolerant individuals. Here, we assessed the number of splenocytes and the cell‐mediated response (proxies for resistance) and body condition (a proxy for tolerance) in four populations of a Greek endemic lizard (Podarcis gaigeae), each exposed to different infection risks (defined as the cumulative effect of parasite burden and duration of exposure). We anticipated that populations with heavy parasite burden would enhance the efficacy of their immune response (resistance) compared to lizards deriving from parasite‐poor habitats. We also predicted that populations with longer exposure to parasites would be adopted and be more tolerant. Each factor (duration of exposure and parasite burden) had a distinct effect on the immune response, and thus, our results were rather complicated. Lizards with heavy parasite burden and aperiodic exposure demonstrated resistance, whereas lizards with heavy parasite burden and chronic exposure were more tolerant. Populations with low parasite burden and minimal exposure were more resistant. Our results suggest that the development of some immunological strategies may be differentiated under different infection risks, even within the same species.     

Long‐term coevolution between avian brood parasites and their hosts

Coevolutionary theory predicts that the most common long‐term outcome of the relationships between brood parasites and their hosts should be coevolutionary cycles based on a dynamic change selecting the currently least‐defended host species, given that when well‐defended hosts are abandoned, hosts will be selected to decrease their defences as these are usually assumed to be costly. This is assumed to be the case also in brood parasite‐host systems. Here I examine the frequency of the three potential long‐term outcomes of brood parasite–host coevolution (coevolutionary cycles, lack of rejection, and successful resistance) in 182 host species. The results of simple exploratory comparisons show that coevolutionary cycles are very scarce while the lack of rejection and successful resistance, which are considered evolutionary enigmas, are much more frequent. I discuss these results considering (i) the importance of different host defences at all stages of the breeding cycle, (ii) the role of phenotypic plasticity in long‐term coevolution, and (iii) the evolutionary history of host selection. I suggest that in purely antagonistic coevolutionary interactions, such as those involving brood parasites and their hosts, that although cycles will exist during an intermediate phase of the interactions, the arms race will end with the extinction of the host or with the host acquiring successful resistance. As evolutionary time passes, this resistance will force brood parasites to use previously less suitable host species. Furthermore, I present a model that represents the long‐term trajectories and outcomes of coevolutionary interactions between brood parasites and their hosts with respect to the evolution of egg‐rejection defence. This model suggests that as an increasing number of species acquire successful resistance, other unparasitized host species become more profitable and their parasitism rate and the costs imposed by brood parasitism at the population level will increase, selecting for the evolution of host defences. This means that although acceptance is adaptive when the parasitism rate and the costs of parasitism are very low, this cannot be considered to represent an evolutionary equilibrium, as conventional theory has done to date, because it is not stable.     

The effect of diet and time after bacterial infection on fecundity,resistance, and tolerance in Drosophila melanogaster

Mounting and maintaining an effective immune response in the face of infection can be costly. The outcome of infection depends on two host immune strategies: resistance and tolerance. Resistance limits pathogen load, while tolerance reduces the fitness impact of an infection. While resistance strategies are well studied, tolerance has received less attention, but is now considered to play a vital role in host–pathogen interactions in animals. A major challenge in ecoimmunology is to understand how some hosts maintain their fitness when infected while others succumb to infection, as well as how extrinsic, environmental factors, such as diet, affect defense. We tested whether dietary restriction through yeast (protein) limitation affects resistance, tolerance, and fecundity in Drosophila melanogaster. We predicted that protein restriction would reveal costs of infection. Because infectious diseases are not always lethal, we tested resistance and tolerance using two bacteria with low lethality: Escherichia coli and Lactococcus lactis. We then assayed fecundity and characterized bacterial infection pathology in individual flies at two acute phase time points after infection. As expected, our four fecundity measures all showed a negative effect of a low‐protein diet, but contrary to predictions, diet did not affect resistance to either bacteria species. We found evidence for diet‐induced and time‐dependent variation in host tolerance to E. coli, but not to L. lactis. Furthermore, the two bacteria species exhibited remarkably different infection profiles, and persisted within the flies for at least 7 days postinfection. Our results show that acute phase infections do not necessarily lead to fecundity costs despite high bacterial loads. The influence of intrinsic variables such as genotype are the prevailing factors that have been studied in relation to variation in host tolerance, but here we show that extrinsic factors should also be considered for their role in influencing tolerance strategies.     

A meta‐analysis of ant social parasitism: host characteristics of different parasitism types and a test of Emery’s rule

Abstract 1. In ant social parasitism, the process by which parasite–host systems evolved and the types of invasion mechanisms parasites use are being debated. Emery’s rule, for example, states that social parasites are the closest relatives to their hosts. The present study uses previously published data to test whether Emery’s rule applies equally to all parasitism types (i.e. xenobiosis, temporary, dulosis, and inquilinism). In addition, this study also investigates other links between parasite–host relatedness and host biology, which has implications for understanding the invasion mechanisms used by certain parasites. 2. We find that xenobiotic parasites typically use distantly‐related host species that are of at least medium colony size. Temporary parasites often have multiple host species that are very closely related to the parasite and hosts with medium‐size colonies. Dulotic parasites frequently have multiple host species that are slightly less related and of any size. Lastly, inquiline parasites tend to have a single, very closely related, host species with medium‐size colonies. 3. Parasites tend to be more closely related to host species if they have a single host species or when the host has a large colony size. In contrast, parasites with multiple host species or hosts of small colony size tend to be less related to their hosts. 4. This study is the first to examine trends in ant social parasitism across all known parasite species. Our meta‐analysis shows that Emery’s rule applies to inquilinism and temporary parasitism, but not to dulosis and xenobiosis. Our results also suggest that both parasitism type and parasite–host relatedness predict the number of hosts and host colony size. It may be that a chemical mimicry mechanism allows invasion of large host colonies, but requires close relatedness of parasite and host, and concentration on a single host species.     

Oviposition preference and larval performance of North American monarch butterflies on four Asclepias species

Monarch butterflies, Danaus plexippus L. (Lepidoptera: Nymphalidae), occur world‐wide and are specialist herbivores of plants in the milkweed family (Asclepiadaceae). In North America, two monarch populations breed east and west of the continental divide in areas populated by different host plant species. To examine the population variation in monarch responses to different Asclepias species, we measured oviposition preference and larval performance among captive progeny reared from adult butterflies collected in eastern and western North America. Host plant use was evaluated using two milkweed species widely distributed in eastern North America (A. incarnata and A. syriaca), and two species common to western North America (A. fascicularis and A. speciosa). We predicted that exposure to different host plant species in their respective breeding ranges could select for divergent host use traits, so that monarchs should preferentially lay more eggs on, and larvae should perform better on, milkweed species common to their native habitats. Results showed that across all adult female butterflies, oviposition preferences were highest for A. incarnata and lowest for A. fascicularis, but mean preferences did not differ significantly between eastern and western monarch populations. Larvae from both populations experienced the highest survival and growth rates on A. incarnata and A. fascicularis, and we again found no significant interactions between monarch source population and milkweed species. Moreover, the average rank order of larval performance did not correspond directly to mean female oviposition preferences, suggesting that additional factors beyond larval performance influence monarch oviposition behavior. Finally, significant family level variation was observed for both preference and performance responses within populations, suggesting an underlying genetic variation or maternal effects governing these traits.     

Host plant species affects virulence in monarch butterfly parasites

1. Studies have considered how intrinsic host and parasite properties determine parasite virulence, but have largely ignored the role of extrinsic ecological factors in its expression. 2. We studied how parasite genotype and host plant species interact to determine virulence of the protozoan parasite Ophryocystis elektroscirrha (McLaughlin & Myers 1970) in the monarch butterfly Danaus plexippus L. We infected monarch larvae with one of four parasite genotypes and reared them on two milkweed species that differed in their levels of cardenolides: toxic chemicals involved in predator defence. 3. Parasite infection, replication and virulence were affected strongly by host plant species. While uninfected monarchs lived equally long on both plant species, infected monarchs suffered a greater reduction in their life spans (55% vs. 30%) on the low-cardenolide vs. the high-cardenolide host plant. These life span differences resulted from different levels of parasite replication in monarchs reared on the two plant species. 4. The virulence rank order of parasite genotypes was unaffected by host plant species, suggesting that host plant species affected parasite genotypes similarly, rather than through complex plant species-parasite genotype interactions. 5. Our results demonstrate that host ecology importantly affects parasite virulence, with implications for host-parasite dynamics in natural populations.     

Evidence for trans-generational medication in nature

Parasites pose a serious threat to host fitness, and natural selection should favour host traits that reduce infection or disease symptoms. Here, we provide the first evidence of trans-generational medication, in which animals actively use medicine to mitigate disease in their offspring. We studied monarch butterflies and their virulent protozoan parasites, and found that neither caterpillars nor adult butterflies could cure themselves of disease. Instead, adult butterflies preferentially laid their eggs on toxic plants that reduced parasite growth and disease in their offspring caterpillars. It has often been suggested that sick animals may use medication to cure themselves of disease, but evidence for the use of medication in nature has so far been scarce. Our results provide evidence that infected animals may indeed use medicine as a defence against parasites, and that such medication may target an individual's offspring rather than the individual itself.     

Virulence evolution in response to anti-infection resistance: toxic food plants can select for virulent parasites of monarch butterflies

Host resistance to parasites can come in two main forms: hosts may either reduce the probability of parasite infection (anti-infection resistance) or reduce parasite growth after infection has occurred (anti-growth resistance). Both resistance mechanisms are often imperfect, meaning that they do not fully prevent or clear infections. Theoretical work has suggested that imperfect anti-growth resistance can select for higher parasite virulence by favouring faster-growing and more virulent parasites that overcome this resistance. In contrast, imperfect anti-infection resistance is thought not to select for increased parasite virulence, because it is assumed that it reduces the number of hosts that become infected, but not the fitness of parasites in successfully infected hosts. Here, we develop a theoretical model to show that anti-infection resistance can in fact select for higher virulence when such resistance reduces the effective parasite dose that enters a host. Our model is based on a monarch butterfly-parasite system in which larval food plants confer resistance to the monarch host. We carried out an experiment and showed that this environmental resistance is most likely a form of anti-infection resistance, through which toxic food plants reduce the effective dose of parasites that initiates an infection. We used these results to build a mathematical model to investigate the evolutionary consequences of food plant-induced resistance. Our model shows that when the effective infectious dose is reduced, parasites can compensate by evolving a higher per-parasite growth rate, and consequently a higher intrinsic virulence. Our results are relevant to many insect host-parasite systems, in which larval food plants often confer imperfect anti-infection resistance. Our results also suggest that - for parasites where the infectious dose affects the within-host dynamics - vaccines that reduce the effective infectious dose can select for increased parasite virulence.     

Vertebrate defense against parasites: Interactions between avoidance,resistance, and tolerance

Hosts can utilize different types of defense against the effects of parasitism, including avoidance, resistance, and tolerance. Typically, there is tremendous heterogeneity among hosts in these defense mechanisms that may be rooted in the costs associated with defense and lead to trade‐offs with other life‐history traits. Trade‐offs may also exist between the defense mechanisms, but the relationships between avoidance, resistance, and tolerance have rarely been studied. Here, we assessed these three defense traits under common garden conditions in a natural host–parasite system, the trematode eye‐fluke Diplostomum pseudospathaceum and its second intermediate fish host. We looked at host individuals originating from four genetically distinct populations of two closely related salmonid species (Atlantic salmon, Salmo salar and sea trout, Salmo trutta trutta) to estimate the magnitude of variation in these defense traits and the relationships among them. We show species‐specific variation in resistance and tolerance and population‐specific variation in resistance. Further, we demonstrate evidence for a trade‐off between resistance and tolerance. Our results suggest that the variation in host defense can at least partly result from a compromise between different interacting defense traits, the relative importance of which is likely to be shaped by environmental components. Overall, this study emphasizes the importance of considering different components of the host defense system when making predictions on the outcome of host–parasite interactions.     

Resistance and tolerance in a host plant-holoparasitic plant interaction: genetic variation and costs

Host organisms are believed to evolve defense mechanisms (i.e., resistance and/or tolerance) under selective pressures exerted by natural enemies. A prerequisite for the evolution of resistance and tolerance is the existence of genetic variation in these traits for natural selection to act. However, selection for resistance and/or tolerance may be constrained by negative genetic correlations with other traits that affect host fitness. We studied genetic variation in resistance and tolerance against parasitic infection and the potential fitness costs associated with these traits using a novel study system, namely the interaction between a flowering plant and a parasitic plant. In this system, parasitic infection has significant negative effects on host growth and reproduction and may thus act as a selective agent. We conducted a greenhouse experiment in which we grew host plants, Urtica dioica, that originated from a single natural population and represented 20 maternal families either uninfected or infected with the holoparasitic dodder, Cuscuta europaea. that originated from the same site. We calculated correlations among resistance, tolerance, and host performance to test for costs of resistance and tolerance. We measured resistance as parasite performance (quantitative resistance) and tolerance as the slopes of regressions relating the vegetative and reproductive biomass of host plants to damage level (measured as parasite biomass). We observed significant differences among host families in parasite resistance and in parasite tolerance in terms of reproductive biomass, a result that suggests genetic variation in these traits. Furthermore, we found differences in resistance and tolerance between female and male host plants. In addition, the correlations indicate costs of resistance in terms of host growth and reproduction and costs of tolerance in terms of host reproduction. Our results thus indicate that host tolerance and resistance can evolve as a response to infection by a parasitic plant and that costs of resistance and tolerance may be one factor maintaining genetic variation in these traits.     

The economy of inflammation: when is less more?

In ecology, tolerance of parasites refers to host mitigation of the fitness costs of an infection. This concept of parasite tolerance contrasts with resistance, whereby hosts reduce the intensity of an infection. Anti-inflammatory cells and molecules have been implicated as mechanisms of parasite tolerance, suggesting that a major role of tolerance is in minimizing collateral damage associated with inflammation. A framework is proposed here in which the cost-benefit outcome of an inflammatory host-response is hypothesized to be dependent on host life-history, parasite virulence, and the efficacy of a current inflammatory or anti-inflammatory response. Testable predictions, both within and among host species, are presented for this hypothesis.     

The evolution of acceptance and tolerance in hosts of avian brood parasites

Avian brood parasites lay their eggs in the nests of their hosts, which rear the parasite's progeny. The costs of parasitism have selected for the evolution of defence strategies in many host species. Most research has focused on resistance strategies, where hosts minimize the number of successful parasitism events using defences such as mobbing of adult brood parasites or rejection of parasite eggs. However, many hosts do not exhibit resistance. Here we explore why some hosts accept parasite eggs in their nests and how this is related to the virulence of the parasite. We also explore the extent to which acceptance of parasites can be explained by the evolution of tolerance; a strategy in which the host accepts the parasite but adjusts its life history or other traits to minimize the costs of parasitism. We review examples of tolerance in hosts of brood parasites (such as modifications to clutch size and multi‐broodedness), and utilize the literature on host–pathogen interactions and plant herbivory to analyse the prevalence of each type of defence (tolerance or resistance) and their evolution. We conclude that (i) the interactions between brood parasites and their hosts provide a highly tractable system for studying the evolution of tolerance, (ii) studies of host defences against brood parasites should investigate both resistance and tolerance, and (iii) tolerance and resistance can lead to contrasting evolutionary scenarios.