Using experimental evolution to explore natural patterns between bacterial motility and resistance to bacteriophages

Resistance of bacteria to phages may be gained by alteration of surface proteins to which phages bind, a mechanism that is likely to be costly as these molecules typically have critical functions such as movement or nutrient uptake. To address this potential trade-off, we combine a systematic study of natural bacteria and phage populations with an experimental evolution approach. We compare motility, growth rate and susceptibility to local phages for 80 bacteria isolated from horse chestnut leaves and, contrary to expectation, find no negative association between resistance to phages and bacterial motility or growth rate. However, because correlational patterns (and their absence) are open to numerous interpretations, we test for any causal association between resistance to phages and bacterial motility using experimental evolution of a subset of bacteria in both the presence and absence of naturally associated phages. Again, we find no clear link between the acquisition of resistance and bacterial motility, suggesting that for these natural bacterial populations, phage-mediated selection is unlikely to shape bacterial motility, a key fitness trait for many bacteria in the phyllosphere. The agreement between the observed natural pattern and the experimental evolution results presented here demonstrates the power of this combined approach for testing evolutionary trade-offs.     

Parasites in bloom: flowers aid dispersal and transmission of pollinator parasites within and between bee species

The dispersal of parasites is critical for epidemiology, and the interspecific vectoring of parasites when species share resources may play an underappreciated role in parasite dispersal. One of the best examples of such a situation is the shared use of flowers by pollinators, but the importance of flowers and interspecific vectoring in the dispersal of pollinator parasites is poorly understood and frequently overlooked. Here, we use an experimental approach to show that during even short foraging periods of 3 h, three bumblebee parasites and two honeybee parasites were dispersed effectively onto flowers by their hosts, and then vectored readily between flowers by non-host pollinator species. The results suggest that flowers are likely to be hotspots for the transmission of pollinator parasites and that considering potential vector, as well as host, species will be of general importance for understanding the distribution and transmission of parasites in the environment and between pollinators.     

Sexual antagonism for resistance and tolerance to infection in Drosophila melanogaster

A critical task in evolutionary genetics is to explain the persistence of heritable variation in fitness-related traits such as immunity. Ecological factors can maintain genetic variation in immunity, but less is known about the role of other factors, such as antagonistic pleiotropy, on immunity. Sexually dimorphic immunity—with females often being more immune-competent—may maintain variation in immunity in dioecious populations. Most eco-immunological studies assess host resistance to parasites rather than the host''s ability to maintain fitness during infection (tolerance). Distinguishing between resistance and tolerance is important as they are thought to have markedly different evolutionary and epidemiological outcomes. Few studies have investigated tolerance in animals, and the extent of sexual dimorphism in tolerance is unknown. Using males and females from 50 Drosophila melanogaster genotypes, we investigated possible sources of genetic variation for immunity by assessing both resistance and tolerance to the common bacterial pathogen Pseudomonas aeruginosa. We found evidence of sexual dimorphism and sexual antagonism for resistance and tolerance, and a trade-off between the two traits. Our findings suggest that antagonistic pleiotropy may be a major contributor to variation in immunity, with implications for host–parasite coevolution.     

The blood parasite Haemoproteus reduces survival in a wild bird: a medication experiment

While avian chronic haemoparasite infections induce reproductive costs, infection has not previously been shown to affect survival. Here, we experimentally reduced, through medication, the intensity of infection by Haemoproteus parasites in wild-breeding female blue tits Cyanistes caeruleus. However, this treatment did not reduce the intensity of infection in males or the intensity of infection by Leucocytozoon. Medicated females, but not males, showed increased local survival until the next breeding season compared with control birds. To our knowledge, this is the first empirical evidence showing long-term direct survival costs of chronic Haemoproteus infections in wild birds.     

Adaptation to abiotic conditions drives local adaptation in bacteria and viruses coevolving in heterogeneous environments

Parasite local adaptation, the greater performance of parasites on their local compared with foreign hosts, has important consequences for the maintenance of diversity and epidemiology. While the abiotic environment may significantly affect local adaptation, most studies to date have failed either to incorporate the effects of the abiotic environment, or to separate them from those of the biotic environment. Here, we tease apart biotic and abiotic components of local adaptation using the bacterium Pseudomonas fluorescens and its viral parasite bacteriophage Φ2. We coevolved replicate populations of bacteria and phages at three different temperatures, and determined their performance against coevolutionary partners from the same and different temperatures. Crucially, we measured performance at different assay temperatures, which allowed us to disentangle adaptation to biotic and abiotic habitat components. Our results show that bacteria and phages are more resistant and infectious, respectively, at the temperature at which they previously coevolved, confirming that local adaptation to abiotic conditions can play a crucial role in determining parasite infectivity and host resistance. Our work underlines the need to assess host–parasite interactions across multiple relevant abiotic environments, and suggests that microbial adaption to local temperatures can create ecological barriers to dispersal across temperature gradients.     

Starvation reveals the cause of infection-induced castration and gigantism

Parasites often induce life-history changes in their hosts. In many cases, these infection-induced life-history changes are driven by changes in the pattern of energy allocation and utilization within the host. Because these processes will affect both host and parasite fitness, it can be challenging to determine who benefits from them. Determining the causes and consequences of infection-induced life-history changes requires the ability to experimentally manipulate life history and a framework for connecting life history to host and parasite fitness. Here, we combine a novel starvation manipulation with energy budget models to provide new insights into castration and gigantism in the Daphnia magna–Pasteuria ramosa host–parasite system. Our results show that starvation primarily affects investment in reproduction, and increasing starvation stress reduces gigantism and parasite fitness without affecting castration. These results are consistent with an energetic structure where the parasite uses growth energy as a resource. This finding gives us new understanding of the role of castration and gigantism in this system, and how life-history variation will affect infection outcome and epidemiological dynamics. The approach of combining targeted life-history manipulations with energy budget models can be adapted to understand life-history changes in other disease systems.     

Disease ecology across soil boundaries: effects of below-ground fungi on above-ground host–parasite interactions

Host–parasite interactions are subject to strong trait-mediated indirect effects from other species. However, it remains unexplored whether such indirect effects may occur across soil boundaries and connect spatially isolated organisms. Here, we demonstrate that, by changing plant (milkweed Asclepias sp.) traits, arbuscular mycorrhizal fungi (AMF) significantly affect interactions between a herbivore (the monarch butterfly Danaus plexippus) and its protozoan parasite (Ophryocystis elektroscirrha), which represents an interaction across four biological kingdoms. In our experiment, AMF affected parasite virulence, host resistance and host tolerance to the parasite. These effects were dependent on both the density of AMF and the identity of milkweed species: AMF indirectly increased disease in monarchs reared on some species, while alleviating disease in monarchs reared on other species. The species-specificity was driven largely by the effects of AMF on both plant primary (phosphorus) and secondary (cardenolides; toxins in milkweeds) traits. Our study demonstrates that trait-mediated indirect effects in disease ecology are extensive, such that below-ground interactions between AMF and plant roots can alter host–parasite interactions above ground. In general, soil biota may play an underappreciated role in the ecology of many terrestrial host–parasite systems.     

Parasites do not adapt to elevated temperature,as evidenced from experimental evolution of a phytoplankton–fungus system

Global warming is predicted to impact the prevalence and severity of infectious diseases. However, empirical data supporting this statement usually stem from experiments in which parasite fitness and disease outcome are measured directly after temperature increase. This might exclude the possibility of parasite adaptation. To incorporate the adaptive response of parasites into predictions of disease severity in a warmer world, we undertook an experimental evolution assay in which a fungal parasite of phytoplankton was maintained at elevated or control temperatures for six months, corresponding to 100–200 parasite generations. Host cultures were maintained at the respective temperatures and provided as substrate, but were not under parasite pressure. A reciprocal infection experiment conducted after six-month serial passages revealed no evidence of parasite adaptation. In fact, parasite fitness at elevated temperatures was inferior in parasite populations reared at elevated temperatures compared with those maintained under control temperature. However, this effect was reversed after parasites were returned to control temperatures for a few (approx. 10) generations. The absence of parasite adaptation to elevated temperatures suggests that, in phytoplankton–fungus systems, disease outcome under global warming will be largely determined by both host and parasite thermal ecology.     

Modularity and specialization in bat–fly interaction networks are remarkably consistent across patches within urbanized landscapes and spatial scales

Patterns of specialization and the structure of interactions between bats and ectoparasitic flies have been studied mostly on non-urban environments and at local scales. Thus, how anthropogenic disturbances influence species interactions and network structure in this system remain poorly understood. Here, we investigated patterns of interaction between Phyllostomidae bats and ectoparasitic Streblidae flies, and variations in network specialization and structure across Cerrado patches within urbanized landscapes in Brazil and between local and regional scales. We found high similarity in the richness and composition of bat and fly species across communities, associated with low turnover of interactions between networks. The high specialization of bat–streblid interactions resulted in little connected and modular networks, with the emergence of modules containing subsets of species that interact exclusively or primarily with each other. Such similarities in species and interaction composition and network structure across communities and scales suggest that bat–fly interactions within Cerrado patches are little affected by the degree of human modification in the surrounding matrix. This remarkable consistency is likely promoted by specific behaviors, the tolerance of Phyllostomidae bats to surrounding urbanized landscapes as well as by the specificity of the streblid–bat interactions shaped over evolutionary time.     

Wildfires modify the parasite loads of invasive cane toads

The frequency and severity of wildfires are increasing due to anthropogenic modifications to habitats and to climate. Post-fire landscapes may advantage invasive species via multiple mechanisms, including changes to host–parasite interactions. We surveyed the incidence of endoparasitic lungworms (Rhabdias pseudosphaerocephala) in invasive cane toads (Rhinella marina) in near-coastal sites of eastern Australia, a year after extensive fires in this region. Both the prevalence of infection and number of worms in infected toads increased with toad body size in unburned areas. By contrast, parasite load decreased with toad body size in burned areas. By killing moisture-dependent free-living lungworm larvae, the intense fires may have liberated adult cane toads from a parasite that can substantially reduce the viability of its host. Smaller toads, which are restricted to moist environments, did not receive this benefit from fires.     

Stochastic environmental fluctuations drive epidemiology in experimental host–parasite metapopulations

Environmental fluctuations are important for parasite spread and persistence. However, the effects of the spatial and temporal structure of environmental fluctuations on host–parasite dynamics are not well understood. Temporal fluctuations can be random but positively autocorrelated, such that the environment is similar to the recent past (red noise), or random and uncorrelated with the past (white noise). We imposed red or white temporal temperature fluctuations on experimental metapopulations of Paramecium caudatum, experiencing an epidemic of the bacterial parasite Holospora undulata. Metapopulations (two subpopulations linked by migration) experienced fluctuations between stressful (5°C) and permissive (23°C) conditions following red or white temporal sequences. Spatial variation in temperature fluctuations was implemented by exposing subpopulations to the same (synchronous temperatures) or different (asynchronous temperatures) temporal sequences. Red noise, compared with white noise, enhanced parasite persistence. Despite this, red noise coupled with asynchronous temperatures allowed infected host populations to maintain sizes equivalent to uninfected populations. It is likely that this occurs because subpopulations in permissive conditions rescue declining subpopulations in stressful conditions. We show how patterns of temporal and spatial environmental fluctuations can impact parasite spread and host population abundance. We conclude that accurate prediction of parasite epidemics may require realistic models of environmental noise.     

Fatal attraction: vegetation responses to nutrient inputs attract herbivores to infectious anthrax carcass sites

Parasites can shape the foraging behaviour of their hosts through cues indicating risk of infection. When cues for risk co-occur with desired traits such as forage quality, individuals face a trade-off between nutrient acquisition and parasite exposure. We evaluated how this trade-off may influence disease transmission in a 3-year experimental study of anthrax in a guild of mammalian herbivores in Etosha National Park, Namibia. At plains zebra (Equus quagga) carcass sites we assessed (i) carcass nutrient effects on soils and grasses, (ii) concentrations of Bacillus anthracis (BA) on grasses and in soils, and (iii) herbivore grazing behaviour, compared with control sites, using motion-sensing camera traps. We found that carcass-mediated nutrient pulses improved soil and vegetation, and that BA is found on grasses up to 2 years after death. Host foraging responses to carcass sites shifted from avoidance to attraction, and ultimately to no preference, with the strength and duration of these behavioural responses varying among herbivore species. Our results demonstrate that animal carcasses alter the environment and attract grazing hosts to parasite aggregations. This attraction may enhance transmission rates, suggesting that hosts are limited in their ability to trade off nutrient intake with parasite avoidance when relying on indirect cues.     

Dispersal network structure and infection mechanism shape diversity in a coevolutionary bacteria-phage system

Resource availability, dispersal and infection genetics all have the potential to fundamentally alter the coevolutionary dynamics of bacteria–bacteriophage interactions. However, it remains unclear how these factors synergise to shape diversity within bacterial populations. We used a combination of laboratory experiments and mathematical modeling to test how the structure of a dispersal network affects host phenotypic diversity in a coevolving bacteria-phage system in communities of differential resource input. Unidirectional dispersal of bacteria and phage from high to low resources consistently increased host diversity compared with a no dispersal regime. Bidirectional dispersal, on the other hand, led to a marked decrease in host diversity. Our mathematical model predicted these opposing outcomes when we incorporated modified gene-for-gene infection genetics. To further test how host diversity depended on the genetic underpinnings of the bacteria-phage interaction, we expanded our mathematical model to include different infection mechanisms. We found that the direction of dispersal had very little impact on bacterial diversity when the bacteria-phage interaction was mediated by matching alleles, gene-for-gene or related infection mechanisms. Our experimental and theoretical results demonstrate that the effects of dispersal on diversity in coevolving host–parasite systems depend on an intricate interplay of the structure of the underlying dispersal network and the specifics of the host–parasite interaction.     

Phylogenetic structure of specialization: A new approach that integrates partner availability and phylogenetic diversity to quantify biotic specialization in ecological networks

Biotic specialization holds information about the assembly, evolution, and stability of biological communities. Partner availabilities can play an important role in enabling species interactions, where uneven partner availabilities can bias estimates of biotic specialization when using phylogenetic diversity indices. It is therefore important to account for partner availability when characterizing biotic specialization using phylogenies. We developed an index, phylogenetic structure of specialization (PSS), that avoids bias from uneven partner availabilities by uncoupling the null models for interaction frequency and phylogenetic distance. We incorporate the deviation between observed and random interaction frequencies as weights into the calculation of partner phylogenetic α‐diversity. To calculate the PSS index, we then compare observed partner phylogenetic α‐diversity to a null distribution generated by randomizing phylogenetic distances among the same number of partners. PSS quantifies the phylogenetic structure (i.e., clustered, overdispersed, or random) of the partners of a focal species. We show with simulations that the PSS index is not correlated with network properties, which allows comparisons across multiple systems. We also implemented PSS on empirical networks of host–parasite, avian seed‐dispersal, lichenized fungi–cyanobacteria, and hummingbird pollination interactions. Across these systems, a large proportion of taxa interact with phylogenetically random partners according to PSS, sometimes to a larger extent than detected with an existing method that does not account for partner availability. We also found that many taxa interact with phylogenetically clustered partners, while taxa with overdispersed partners were rare. We argue that species with phylogenetically overdispersed partners have often been misinterpreted as generalists when they should be considered specialists. Our results highlight the important role of randomness in shaping interaction networks, even in highly intimate symbioses, and provide a much‐needed quantitative framework to assess the role that evolutionary history and symbiotic specialization play in shaping patterns of biodiversity. PSS is available as an R package at https://github.com/cjpardodelahoz/pss.     

Mothers produce less aggressive sons with altered immunity when there is a threat of disease during pregnancy

Maternal experience before and during pregnancy is known to play a key role in offspring development. However, the influence of social cues about disease in the maternal environment has not been explored. We indirectly exposed pregnant mice to infected neighbours by housing them next to non-contagious conspecifics infected with Babesia microti. We examined the effect of this indirect immunological exposure on both the females and their adult offspring. Exposed females had higher levels of serum corticosterone and increased kidney growth compared with those with uninfected neighbours. These exposed females subsequently produced offspring that as adults showed an accelerated immune response to B. microti and less aggression in social groups. We suggest that ambient information regarding disease is used adaptively to maximize offspring survival and reproductive success in a challenging environment. Our results shed light on the impact of social information and maternal effects on life histories, and have important consequences for our understanding of epidemiology and individual disease susceptibility in humans and other animals. They also lead us to question the suitability of some laboratory housing conditions during experimental procedures, which may impact negatively upon both animal welfare and the validity of animal science.     

Sea lice and salmon population dynamics: effects of exposure time for migratory fish

The ecological impact of parasite transmission from fish farms is probably mediated by the migration of wild fishes, which determines the period of exposure to parasites. For Pacific salmon and the parasitic sea louse, Lepeophtheirus salmonis, analysis of the exposure period may resolve conflicting observations of epizootic mortality in field studies and parasite rejection in experiments. This is because exposure periods can differ by 2–3 orders of magnitude, ranging from months in the field to hours in experiments. We developed a mathematical model of salmon–louse population dynamics, parametrized by a study that monitored naturally infected juvenile salmon held in ocean enclosures. Analysis of replicated trials indicates that lice suffer high mortality, particularly during pre-adult stages. The model suggests louse populations rapidly decline following brief exposure of juvenile salmon, similar to laboratory study designs and data. However, when the exposure period lasts for several weeks, as occurs when juvenile salmon migrate past salmon farms, the model predicts that lice accumulate to abundances that can elevate salmon mortality and depress salmon populations. The duration of parasite exposure is probably critical to salmon–louse population dynamics, and should therefore be accommodated in coastal planning and management where fish farms are situated on wild fish migration routes.     

Rapidly fluctuating environments constrain coevolutionary arms races by impeding selective sweeps

Although pervasive, the impact of temporal environmental heterogeneity on coevolutionary processes is poorly understood. Productivity is a key temporally heterogeneous variable, and increasing productivity has been shown to increase rates of antagonistic arms race coevolution, and lead to the evolution of more broadly resistant hosts and more broadly infectious parasites. We investigated the effects of the grain of environmental heterogeneity, in terms of fluctuations in productivity, on bacteria–phage coevolution. Our findings demonstrate that environmental heterogeneity could constrain antagonistic coevolution, but that its effect was dependent upon the grain of heterogeneity, such that both the rate and extent of coevolution were most strongly limited in fine-grained, rapidly fluctuating heterogeneous environments. We further demonstrate that rapid environmental fluctuations were likely to have impeded selective sweeps of resistance alleles, which occurred over longer durations than the fastest, but not the slowest, frequency of fluctuations used. Taken together our results suggest that fine-grained environmental heterogeneity constrained the coevolutionary arms race by impeding selective sweeps.     

Females that experience threat are better teachers

Superb fairy-wren (Malurus cyaneus) females use an incubation call to teach their embryos a vocal password to solicit parental feeding care after hatching. We previously showed that high call rate by the female was correlated with high call similarity in fairy-wren chicks, but not in cuckoo chicks, and that parent birds more often fed chicks with high call similarity. Hosts should be selected to increase their defence behaviour when the risk of brood parasitism is highest, such as when cuckoos are present in the area. Therefore, we experimentally test whether hosts increase call rate to embryos in the presence of a singing Horsfield''s bronze-cuckoo (Chalcites basalis). Female fairy-wrens increased incubation call rate when we experimentally broadcast cuckoo song near the nest. Embryos had higher call similarity when females had higher incubation call rate. We interpret the findings of increased call rate as increased teaching effort in response to a signal of threat.     

Antagonistic coevolution with parasites maintains host genetic diversity: an experimental test

Genetic variation in natural populations is a prime prerequisite allowing populations to respond to selection, but is under constant threat from forces that tend to reduce it, such as genetic drift and many types of selection. Haldane emphasized the potential importance of parasites as a driving force of genetic diversity. His theory has been taken for granted ever since, but despite numerous studies showing correlations between genetic diversity and parasitism, Haldane''s hypothesis has rarely been tested experimentally for unambiguous support. We experimentally staged antagonistic coevolution between the host Tribolium castaneum and its natural microsporidian parasite, Nosema whitei, to test for the relative importance of two separate evolutionary forces (drift and parasite-induced selection) on the maintenance of genetic variation. Our results demonstrate that coevolution with parasites indeed counteracts drift as coevolving populations had significantly higher levels of heterozygosity and allelic diversity. Genetic drift remained a strong force, strongly reducing genetic variation and increasing genetic differentiation in small populations. To our surprise, differentiation between the evolving populations was smaller when they coevolved with parasites, suggesting parallel balancing selection. Hence, our results experimentally vindicate Haldane''s original hypothesis 60 years after its conception.