Host Defense Mechanisms Induce Genome Instability Leading to Rapid Evolution in an Opportunistic Fungal Pathogen.

The ability to generate genetic variation facilitates rapid adaptation in stressful environments. The opportunistic fungal pathogen Candida albicans frequently undergoes large-scale genomic changes, including aneuploidy and loss of heterozygosity (LOH), following exposure to host environments. However, the specific host factors inducing C. albicans genome instability remain largely unknown. Here, we leveraged the genetic tractability of nematode hosts to investigate whether innate immune components, including antimicrobial peptides (AMPs) and reactive oxygen species (ROS), induced host-associated C. albicans genome instability. C. albicans associated with immunocompetent hosts carried multiple large-scale genomic changes, including LOH and whole-chromosomal and segmental aneuploidies. In contrast, C. albicans associated with immunocompromised hosts deficient in AMPs or ROS production had reduced LOH frequencies and fewer, if any, additional genomic changes. To evaluate whether extensive host-induced genomic changes had long-term consequences for C. albicans adaptation, we experimentally evolved C. albicans in either immunocompetent or immunocompromised hosts and selected for increased virulence. C. albicans evolved in immunocompetent hosts rapidly increased virulence, but C. albicans evolved in immunocompromised hosts did not. Taken together, this work suggests that host-produced ROS and AMPs induces genotypic plasticity in C. albicans which facilitates rapid evolution.

Advantages of laboratory natural selection in the applied sciences.

In the past three decades, laboratory natural selection has become a widely used technique in biological research. Most studies which have utilized this technique are in the realm of basic science, often testing hypotheses related to mechanisms of evolutionary change or ecological dynamics. While laboratory natural selection is currently utilized heavily in this setting, there is a significant gap with its usage in applied studies, especially when compared to the other selection experiment methodologies like artificial selection and directed evolution. This is despite avenues of research in the applied sciences which seem well suited to laboratory natural selection. In this review, we place laboratory natural selection in context with other selection experiments, identify the characteristics which make it well suited for particular kinds of applied research and briefly cover key examples of the usefulness of selection experiments within applied science. Finally, we identify three promising areas of inquiry for laboratory natural selection in the applied sciences: bioremediation technology, identifying mechanisms of drug resistance and optimizing biofuel production. Although laboratory natural selection is currently less utilized in applied science when compared to basic research, the method has immense promise in the field moving forward.

A need to consider the evolutionary genetics of host-symbiont mutualisms.

Despite the ubiquity and importance of mutualistic interactions, we know little about the evolutionary genetics underlying their long-term persistence. As in antagonistic interactions, mutualistic symbioses are characterized by substantial levels of phenotypic and genetic diversity. In contrast to antagonistic interactions, however, we, by and large, do not understand how this variation arises, how it is maintained, nor its implications for future evolutionary change. Currently, we rely on phenotypic models to address the persistence of mutualistic symbioses, but the success of an interaction almost certainly depends heavily on genetic interactions. In this review, we argue that evolutionary genetic models could provide a framework for understanding the causes and consequences of diversity and why selection may favour processes that maintain variation in mutualistic interactions.

The evolution of parasite host range in heterogeneous host populations.

Theory on the evolution of niche width argues that resource heterogeneity selects for niche breadth. For parasites, this theory predicts that parasite populations will evolve, or maintain, broader host ranges when selected in genetically diverse host populations relative to homogeneous host populations. To test this prediction, we selected the bacterial parasite Serratia marcescens to kill Caenorhabditis elegans in populations that were genetically heterogeneous (50% mix of two experimental genotypes) or homogeneous (100% of either genotype). After 20 rounds of selection, we compared the host range of selected parasites by measuring parasite fitness (i.e. virulence, the selected fitness trait) on the two focal host genotypes and on a novel host genotype. As predicted, heterogeneous host populations selected for parasites with a broader host range: these parasite populations gained or maintained virulence on all host genotypes. This result contrasted with selection in homogeneous populations of one host genotype. Here, host range contracted, with parasite populations gaining virulence on the focal host genotype and losing virulence on the novel host genotype. This pattern was not, however, repeated with selection in homogeneous populations of the second host genotype: these parasite populations did not gain virulence on the focal host genotype, nor did they lose virulence on the novel host genotype. Our results indicate that host heterogeneity can maintain broader host ranges in parasite populations. Individual host genotypes, however, vary in the degree to which they select for specialization in parasite populations.

Evolution of increased virulence is associated with decreased spite in the insect-pathogenic bacterium Xenorhabdus nematophila.

Disease virulence may be strongly influenced by social interactions among pathogens, both during the time course of an infection and evolutionarily. Here, we examine how spiteful bacteriocin production in the insect-pathogenic bacterium Xenorhabdus nematophila is evolutionarily linked to its virulence. We expected a negative correlation between virulence and spite owing to their inverse correlations with growth. We examined bacteriocin production and growth across 14 experimentally evolved lineages that show faster host-killing relative to their ancestral population. Consistent with expectations, these more virulent lineages showed reduced bacteriocin production and faster growth relative to the ancestor. Further, bacteriocin production was negatively correlated with growth across the examined lineages. These results strongly support an evolutionary trade-off between virulence and bacteriocin production and lend credence to the view that disease management can be improved by exploiting pathogen social interactions.

No measurable fitness cost to experimentally evolved host defence in the Caenorhabditis elegans-Serratia marcescens host-parasite system.

Host susceptibility to parasites can vary over space and time. Costs associated with the maintenance of host defence are thought to account for a portion of this variation. Specifically, trade-offs wherein elevated defence is maintained at the cost of fitness in the absence of the parasite may cause levels of host defence to change over time and differ between populations. In previous studies, we found that populations of the host nematode, Caenorhabditis elegans, evolved greater levels of parasite avoidance and resistance against the bacterial parasite, Serratia marcescens. Here, we passaged these host populations either in the presence or absence of the parasite to test for a cost of elevated host defences. After 16 generations, we found that elevated levels of host defence were maintained during evolution in both the presence and absence of the parasite. Further, this maintenance of defence was not the result of limited standing genetic variation, but rather the absence of a measurable cost associated with defence. Therefore, costs associated with host defence may not broadly account for differences in host susceptibility across space and time.

Restoration of pyrethroid susceptibility in a highly resistant Aedes aegypti population.

Insecticide resistance has evolved in disease vectors worldwide, creating the urgent need to either develop new control methods or restore insecticide susceptibility to regain use of existing tools. Here we show that phenotypic susceptibility can be restored in a highly resistant field-derived strain of Aedes aegypti in only 10 generations through rearing them in the absence of insecticide.

Host mating system and coevolutionary dynamics shape the evolution of parasite avoidance in Caenorhabditis elegans host populations.

Hosts exhibit a variety of defence mechanisms against parasites, including avoidance. Both host-parasite coevolutionary dynamics and the host mating system can alter the evolutionary trajectories of populations. Does the nature of host-parasite interactions and the host mating system affect the mechanisms that evolve to confer host defence? In a previous experimental evolution study, mixed mating and obligately outcrossing Caenorhabditis elegans host populations adapted to either coevolving or static Serratia marcescens parasite populations. Here, we assessed parasite avoidance as a mechanism underlying host adaptation. We measured host feeding preference for the coevolved and static parasites vs preference for Escherichia coli, to assess the evolution of avoidance behaviour within our experiment. We found that mixed mating host populations evolved a preference for E. coli relative to the static parasite strain; therefore, the hosts evolved parasite avoidance as a defence. However, mixed mating hosts did not exhibit E. coli preference when exposed to coevolved parasites, so avoidance cannot account for host adaptation to coevolving parasites. Further, the obligately outcrossing host populations did not exhibit parasite avoidance in the presence of either static or coevolved parasites. Therefore, both the nature of host-parasite interactions and the host mating system shaped the evolution of host defence.

A Model for Evolutionary Ecology of Disease: The Case for Caenorhabditis Nematodes and Their Natural Parasites.

Many of the outstanding questions in disease ecology and evolution call for combining observation of natural host-parasite populations with experimental dissection of interactions in the field and the laboratory. The "rewilding" of model systems holds great promise for this endeavor. Here, we highlight the potential for development of the nematode Caenorhabditis elegans and its close relatives as a model for the study of disease ecology and evolution. This powerful laboratory model was disassociated from its natural habitat in the 1960s. Today, studies are uncovering that lost natural history, with several natural parasites described since 2008. Studies of these natural Caenorhabditis-parasite interactions can reap the benefits of the vast array of experimental and genetic tools developed for this laboratory model. In this review, we introduce the natural parasites of C. elegans characterized thus far and discuss resources available to study them, including experimental (co)evolution, cryopreservation, behavioral assays, and genomic tools. Throughout, we present avenues of research that are interesting and feasible to address with caenorhabditid nematodes and their natural parasites, ranging from the maintenance of outcrossing to the community dynamics of host-associated microbes. In combining natural relevance with the experimental power of a laboratory supermodel, these fledgling host-parasite systems can take on fundamental questions in evolutionary ecology of disease.

Mutation load and rapid adaptation favour outcrossing over self-fertilization.

The tendency of organisms to reproduce by cross-fertilization despite numerous disadvantages relative to self-fertilization is one of the oldest puzzles in evolutionary biology. For many species, the primary obstacle to the evolution of outcrossing is the cost of production of males, individuals that do not directly contribute offspring and thus diminish the long-term reproductive output of a lineage. Self-fertilizing ('selfing') organisms do not incur the cost of males and therefore should possess at least a twofold numerical advantage over most outcrossing organisms. Two competing explanations for the widespread prevalence of outcrossing in nature despite this inherent disadvantage are the avoidance of inbreeding depression generated by selfing and the ability of outcrossing populations to adapt more rapidly to environmental change. Here we show that outcrossing is favoured in populations of Caenorhabditis elegans subject to experimental evolution both under conditions of increased mutation rate and during adaptation to a novel environment. In general, fitness increased with increasing rates of outcrossing. Thus, each of the standard explanations for the maintenance of outcrossing are correct, and it is likely that outcrossing is the predominant mode of reproduction in most species because it is favoured under ecological conditions that are ubiquitous in natural environments.

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

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

Outcrossing increases resistance against coevolving parasites.

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

Outcrossing in Caenorhabditis elegans increases in response to food limitation.

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

Gene flow accelerates adaptation to a parasite.

Gene flow into populations can increase additive genetic variation and introduce novel beneficial alleles, thus facilitating adaptation. However, gene flow may also impede adaptation by disrupting beneficial genotypes, introducing deleterious alleles, or creating novel dominant negative interactions. While theory and fieldwork have provided insight into the effects of gene flow, direct experimental tests are rare. Here, we evaluated the effects of gene flow on adaptation in the nematode Caenorhabditis elegans during exposure to the bacterial parasite, Serratia marcescens. We evolved hosts against nonevolving parasites for 10 passages while controlling host gene flow and source population. We used source nematode populations with three different genetic backgrounds (one similar to the sink population and two different) and two evolutionary histories (previously adapted to S. marcescens or naive). We found that populations with gene flow exhibited greater increases in parasite resistance than those without gene flow. Additionally, gene flow from adapted populations resulted in greater increases in resistance than gene flow from naive populations, particularly with gene flow from novel genetic backgrounds. Overall, this work demonstrates that gene flow can facilitate adaptation and suggests that the genetic architecture and evolutionary history of source populations can alter the sink population's response to selection.

Evaluating coevolution in a horizontally transmitted mutualism.

Many interspecific interactions are shaped by coevolution. Transmission mode is thought to influence opportunities for coevolution within symbiotic interactions. Vertical transmission maintains partner fidelity, increasing opportunities for coevolution, but horizontal transmission may disrupt partner fidelity, potentially reducing opportunities for coevolution. Despite these predictions, the role of coevolution in the maintenance of horizontally transmitted symbioses is unclear. Leveraging a tractable insect-bacteria symbiosis, we tested for signatures of pairwise coevolution by assessing patterns of host-symbiont specialization. If pairwise coevolution defines the interaction, we expected to observe evidence of reciprocal specialization between hosts and their local symbionts. We found no evidence for local adaptation between sympatric lineages of Anasa tristis squash bugs and Caballeronia spp. symbionts across their native geographic range. We also found no evidence for specialization between three co-localized Anasa host species and their native Caballeronia symbionts. Our results demonstrate generalist dynamics underlie the interaction between Anasa insect hosts and their Caballeronia symbionts. We predict that selection from multiple host species may favor generalist symbiont traits through diffuse coevolution. Alternatively, selection for generalist traits may be a consequence of selection by hosts for fixed cooperative symbiont traits without coevolution.

High parasite virulence necessary for the maintenance of host outcrossing via parasite-mediated selection.

Biparental sex is widespread in nature, yet costly relative to uniparental reproduction. It is generally unclear why self-fertilizing or asexual lineages do not readily invade outcrossing populations. The Red Queen hypothesis predicts that coevolving parasites can prevent self-fertilizing or asexual lineages from invading outcrossing host populations. However, only highly virulent parasites are predicted to maintain outcrossing, which may limit the general applicability of the Red Queen hypothesis. Here, we tested whether the ability of coevolving parasites to prevent invasion of self-fertilization within outcrossing host populations was dependent on parasite virulence. We introduced wild-type Caenorhabditis elegans hermaphrodites, capable of both self-fertilization and outcrossing, into C. elegans populations fixed for a mutant allele conferring obligate outcrossing. Replicate C. elegans populations were exposed for 24 host generations to one of four strains of Serratia marcescens parasites that varied in virulence, under three treatments: a heat-killed (control, noninfectious) parasite treatment, a fixed-genotype (nonevolving) parasite treatment, and a copassaged (potentially coevolving) parasite treatment. As predicted, self-fertilization invaded C. elegans host populations in the control and fixed-parasite treatments, regardless of parasite virulence. In the copassaged treatment, selfing invaded host populations coevolving with low- to mid-virulence strains, but remained rare in hosts coevolving with highly virulent bacterial strains. Therefore, we found that only highly virulent coevolving parasites can impede the invasion of selfing.

Host-associated transmission favors transition of a commensal toward antagonism.

The impacts of host-associated microbes on their hosts vary along a continuum of antagonistic, neutral, and beneficial interactions. Transmission mode is predicted to contribute to transitions along the continuum by altering opportunities for the alignment of host and microbe fitness interests. Under vertical transmission, microbial evolution is tightly coupled to the host environment, which may facilitate fitness alignment. In contrast, environmentally transmitted microbes spend time in the external environment, outside of hosts, partially decoupling their evolution from the host. This decoupling may misalign host and microbe fitness interests, potentially favoring antagonistic microbial traits. Here, we tested whether transmission environment alters microbial evolution by manipulating the interaction between a commensal Serratia marcescens bacteria and their insect host Anasa tristis, which is the primary vector of these bacteria into plants, where they cause disease. We experimentally evolved S. marcescens through several selection environments. The bacteria were passaged between A. tristis hosts, between A. tristis hosts and soil, through soil, or through standard culture media. We observed rapid evolution of virulence toward hosts across treatments when bacterial evolution occurred within the host environment, indicating that direct host-to-host transmission can increase opportunities for microbes to adapt to hosts and evolve antagonistic traits.

Coevolution's conflicting role in the establishment of beneficial associations.

Reciprocal adaptation between hosts and symbionts can drive the maintenance of symbioses, resulting in coevolution and beneficial genotypic interactions. Consequently, hosts may experience decreased fitness when paired with nonsympatric partners compared to sympatric symbionts. However, coevolution does not preclude conflict-host and symbiont can act to advance their own fitness interests, which do not necessarily align with those of their partner. Despite coevolution's importance in extant symbioses, we know little about its role in shaping the origin of symbioses. Here, we tested the role of coevolution in establishing a novel association by experimentally (co)evolving a host with a protective bacterium under environmental stress. Although evolution in the presence of nonevolving bacteria facilitated host adaptation, co-passaged hosts did not exhibit greater adaptation rates than hosts paired with nonevolving bacteria. Furthermore, co-passaged hosts exhibited greater fecundity when paired with sympatric, co-passaged bacteria compared to co-passaged bacteria with which they did not share an evolutionary history. Thus, shared evolutionary history between the hosts and microbes actually reduced host fitness and has the potential to impede evolution of new beneficial associations.

Increased Virulence and Large-Scale Reduction in Genome Size of Tetraploid Candida albicans Evolved in Nematode Hosts.

Candida albicans is an opportunistic fungal pathogen of humans, yet the within-host dynamics of C. albicans infection are not clear. While C. albicans is commonly diploid, it exhibits a range of ploidies, including tetraploidy. Previous work found that tetraploid C. albicans populations exhibited rapid adaptation and significant genome instability when evolved in vitro. Host immune function alters the rate and magnitude of C. albicans virulence evolution, but the effects of the host immunity on tetraploid C. albicans populations are unclear. Here, we tested the effects of the host immunity on genome stability and virulence evolution of tetraploid C. albicans using experimental evolution. We selected for C. albicans increased virulence within either immunocompetent or immunocompromised Caenorhabditis elegans hosts. After nine passages we observed a response to selection for increased virulence. Both populations exposed to either immunocompetent or immunocompromised hosts increased virulence after passage through C. elegans hosts. However, the C. albicans populations passaged through immunocompetent hosts under selection exhibited unique temporal dynamics, a rapid increase in virulence and then subsequent loss of virulence. Most C. albicans populations exhibited genome size reduction within six passages, however populations exposed to immunocompetent hosts exhibited the most rapid transition to ~diploid. Therefore, we found that tetraploids rapidly increase in virulence and decrease genome size within host environments. Further, the combination of selection for greater virulence in the presence of immunocompetent hosts results in major virulence fluctuations and genome size changes. Thus, host immunity significantly impacts the evolutionary trajectories of tetraploid C. albicans.

Association with a novel protective microbe facilitates host adaptation to a stressful environment.

Protective symbionts can allow hosts to occupy otherwise uninhabitable niches. Despite the importance of symbionts in host evolution, we know little about how these associations arise. Encountering a microbe that can improve host fitness in a stressful environment may favor persistent interactions with that microbe, potentially facilitating a long-term association. The bacterium Bacillus subtilis protects Caenorhabditis elegans nematodes from heat shock by increasing host fecundity compared to the nonprotective Escherichia coli. In this study, we ask how the protection provided by the bacterium affects the host's evolutionary trajectory. Because of the stark fitness contrast between hosts heat shocked on B. subtilis versus E. coli, we tested whether the protection conferred by the bacteria could increase the rate of host adaptation to a stressful environment. We passaged nematodes on B. subtilis or E. coli, under heat stress or standard conditions for 20 host generations of selection. When assayed under heat stress, we found that hosts exhibited the greatest fitness increase when evolved with B. subtilis under stress compared to when evolved with E. coli or under standard (nonstressful) conditions. Furthermore, despite not directly selecting for increased B. subtilis fitness, we found that hosts evolved to harbor more B. subtilis as they adapted to heat stress. Our findings demonstrate that the context under which hosts evolve is important for the evolution of beneficial associations and that protective microbes can facilitate host adaptation to stress. In turn, such host adaptation can benefit the microbe.