Natural variation in priming of basal resistance: from evolutionary origin to agricultural exploitation

Biotic stress has a major impact on the process of natural selection in plants. As plants have evolved under variable environmental conditions, they have acquired a diverse spectrum of defensive strategies against pathogens and herbivores. Genetic variation in the expression of plant defence offers valuable insights into the evolution of these strategies. The 'zigzag' model, which describes an ongoing arms race between inducible plant defences and their suppression by pathogens, is now a commonly accepted model of plant defence evolution. This review explores additional strategies by which plants have evolved to cope with biotic stress under different selective circumstances. Apart from interactions with plant-beneficial micro-organisms that can antagonize pathogens directly, plants have the ability to prime their immune system in response to selected environmental signals. This defence priming offers disease protection that is effective against a broad spectrum of virulent pathogens, as long as the augmented defence reaction is expressed before the invading pathogen has the opportunity to suppress host defences. Furthermore, priming has been shown to be a cost-efficient defence strategy under relatively hostile environmental conditions. Accordingly, it is possible that selected plant varieties have evolved a constitutively primed immune system to adapt to levels of disease pressure. Here, we examine this hypothesis further by evaluating the evidence for natural variation in the responsiveness of basal defence mechanisms, and discuss how this genetic variation can be exploited in breeding programmes to provide sustainable crop protection against pests and diseases.     

Does ant predation favour leaf beetle specialization on toxic host plants?

Generalist predators are frequently seen as evolutionary forces that narrow the host range in herbivorous insects. Predators may favour specialization of herbivores on host plants containing toxic chemicals (which can be used by herbivores for their own defence) if host plant‐derived defences provide better protection from enemies than do autogenously produced defences. We compared the effectiveness of these two defensive strategies in the larvae of six species of leaf beetle (Chrysomelidae) against wood ants (Formica rufa group) in field experiments. Ants were more strongly repelled by larvae with host plant‐derived, salicylaldehyde‐containing secretions than by larvae with various autogenous secretions, but collectively foraging ants ultimately overcame any type of chemical defence by social interactions, chemical signalling, and olfactory learning. As a result, ants killed all larvae of Chrysomela lapponica defended by salicylaldehyde‐containing secretions within 2 days of their introduction to willows within 15 m of ant nests. We conclude that in the field neither type of chemical defence provides complete protection against wood ants in the vicinity of their nests, and that evolutionary shifts from autogenous production of secretion to sequestration of plant allelochemicals in leaf beetles may be favoured mostly at low ant densities on the periphery of ant foraging areas.     

Strategies of chemical anti-predator defences in leaf beetles: is sequestration of plant toxins less costly than de novo synthesis?

The evolution of defensive traits is driven both by benefits gained from protection against enemies and by costs of defence production. We tested the hypothesis that specialisation of herbivores on toxic host plants, accompanied by the ability to acquire plant defensive compounds for herbivore defence, is favoured by the lower costs of sequestration compared to de novo synthesis of defensive compounds. We measured physiological costs of chemical defence as a reduction in larval performance in response to repeated removal of secretions (simulating predator attack) and compared these costs between five species synthesising defences de novo and three species sequestering salicylic glucosides (SGs) from their host plants. Experiments simulating low predator pressure revealed no physiological costs in terms of survival, weight and duration of development in any of study species. However, simulation of high predation caused reduction in relative growth rate in Chrysomela lapponica larvae producing autogenous defences more frequently, than in larvae sequestering SGs. Still meta-analysis of combined data showed no overall difference in costs of autogenous and sequestered defences. However, larvae synthesising their defences de novo demonstrated secretion-conserving behaviour, produced smaller amounts of secretions, replenished them at considerably lower rates and employed other types of defences (regurgitation, evasion) more frequently when compared to sequestering larvae. These latter results provide indirect evidence for biosynthetic constraints for amounts of defensive secretions produced de novo, resulting in low defence effectiveness. Lifting these constraints by sequestration may have driven some leaf beetle lineages toward sequestration of plant allelochemicals as the main defensive strategy.     

Divergent immune priming responses across flour beetle life stages and populations

Growing evidence shows that low doses of pathogens may prime the immune response in many insects, conferring subsequent protection against infection in the same developmental stage (within‐life stage priming), across life stages (ontogenic priming), or to offspring (transgenerational priming). Recent work also suggests that immune priming is a costly response. Thus, depending on host and pathogen ecology and evolutionary history, tradeoffs with other fitness components may constrain the evolution of priming. However, the relative impacts of priming at different life stages and across natural populations remain unknown. We quantified immune priming responses of 10 natural populations of the red flour beetle Tribolium castaneum, primed and infected with the natural insect pathogen Bacillus thuringiensis. We found that priming responses were highly variable both across life stages and populations, ranging from no detectable response to a 13‐fold survival benefit. Comparing across stages, we found that ontogenic immune priming at the larval stage conferred maximum protection against infection. Finally, we found that various forms of priming showed sex‐specific associations that may represent tradeoffs or shared mechanisms. These results indicate the importance of sex‐, life stage‐, and population‐specific selective pressures that can cause substantial divergence in priming responses even within a species. Our work highlights the necessity of further work to understand the mechanistic basis of this variability.     

Galapagos ground finches balance investment in behavioural and immunological pathogen defences

The scientific community has long recognized the importance of individual behaviour in pathogen spread in wild animals. Furthermore, recent studies have highlighted the potential for behaviour to act as a pathogen defence mechanism. In line with these studies, the pathogen defence optimization hypothesis (PDOH) posits that individuals balance investment in costly behavioural and immunological defences against pathogen infection. Here, I test the PDOH using data from observations of wild Galapagos finches (Medium Ground Finches Geospiza fortis and Small Ground Finches Geospiza fuliginosa) at feeders, combined with immune data collected in the field. Both within and between species, those groups engaging to a greater extent in high transmission‐risk behaviours (not investing in behavioural defences against pathogen exposure) invest more heavily in immune pathogen defences. These data provide the first support for the PDOH in a wild population.     

Physical and chemical properties of primary defences in Tenebrio molitor

Prevention and reaction are the foundation for any defence system. In insects, the primary defences against pathogens and parasites limit invasion; the secondary ones (e.g. immune system) act when the cuticle and other primary defences fail. Because investment in both aspects of defence may be costly, they should be regulated in a plastic or variable way in accordance with the risk of infection. The mealworm beetle Tenebrio molitor L. changes cuticle colour and its resistance to fungal infection when subject to high population density, although such resistance is a result of the primary (cuticle) defences rather than the secondary (immunological) ones. The present study tests the hypothesis that the physical and chemical properties of the primary defences in T. molitor change with cuticular darkness. Beetles expressing black phenotypes (or with darker cuticle) have a thicker cuticle, with four well organized layers (epi‐, exo‐, endocuticle and formation zone) and more melanin than tan beetles. The cuticle properties investigated in the present study are likely to be the underlying mechanisms of pathogen resistance in black beetles, including the content of carbonylated proteins, which in black beetles was almost half that of tan beetles after exposure to ultraviolet radiation. It is proposed that, in polyphenic insects (such as mealworm beetles), primary and secondary defences are regulated pleiotropically, with the genes responsible for the expression of one defence having a positive effect on others, whereas, in polymorphic insects, there is no such link and so investment in one defence may impair others.     

The Lactoperoxidase System: a Natural Biochemical Biocontrol Agent for Pre‐ and Postharvest Applications

Controlling pests in pre‐ and postharvest crops using natural and low‐impact products is a major challenge. The lactoperoxidase system is an enzymatic system that exists in all external secretions in mammals and is part of the non‐immune system. We tested its efficacy in in vitro microplates on Phytophthora infestans, Penicillium digitatum, Penicillium italicum, Penicillium expansum and Botrytis cinerea to determine the most suitable concentrations for use. Then, we verified its efficacy in planta under controlled conditions. Solutions prepared with 5.4 mm iodide and 1.2 mm thiocyanate and diluted threefold inhibited pathogen growth in vitro by 63–100%. Twofold‐diluted solutions protected potato plants against P. infestans by 60–74% under controlled conditions. Undiluted solution inhibited orange's and apple's postharvest pathogens in curative application with efficacy levels ranging between 84 and 95% in orange and between 63 and 74% in apple. 1.5‐fold concentrated solutions inhibited postharvest pathogens of apple in curative application with efficacy levels ranging between 84 and 92%. Our results also show that the oxidative stress response of fruit following wounding could interfere with ion efficiency. Our tests demonstrate for the first time that this biochemical method is as efficient as a conventional synthetic chemical method under controlled conditions.     

Do pygidial secretions of dung beetles have the potential to repel urban pest ants?

Various organisms emit malodorous secretions against competitors, and the potential use of these secretions in pest management should be investigated. For example, some ant species feed on similar resources as dung beetles, which might have led to counter chemical defences in dung beetles. We tested whether pygidial secretions of the dung beetle Canthon smaragdulus (Fabricius) (Coleoptera: Scarabaeidae, Scarabaeinae) alter the locomotor behaviour of the exotic urban pest ant Tapinoma melanocephalum (Fabricius) (Hymenoptera: Formicidae), specifically whether these secretions repel those ants. We also tested whether the disturbance in the locomotor behaviour of T. melanocephalum increases with the amount of pygidial secretion. We found that individual T. melanocephalum displayed changes in their locomotor behaviour when exposed to pygidial secretions of coupled dung beetles, single males, and single females. Additionally, the pygidial secretions from male and female dung beetles could repel ants. The change in the locomotor behaviour of T. melanocephalum increased with the amount of pygidial secretion. Our results suggest that the pygidial secretions of dung beetles have potential as a biological repellent of T. melanocephalum. Hence, pygidial secretions from dung beetles may be used in the future for the development of urban pest management strategies.     

Defence against multiple enemies

Although very common under natural conditions, the consequences of multiple enemies (parasites, predators, herbivores, or even 'chemical' enemies like insecticides) on investment in defence has scarcely been investigated. In this paper, we present a simple model of the joint evolution of two defences targeted against two enemies. We illustrate how the respective level of each defence can be influenced by the presence of the two enemies. Furthermore, we investigate the influences of direct interference and synergy between defences. We show that, depending on certain conditions (costs, interference or synergy between defences), an increase in selection pressure by one enemy can have dramatic effects on defence against another enemy. It is generally admitted that increasing the encounter rate with a second natural enemy can decrease investment in defence against a first enemy, but our results indicate that it may sometimes favour resistance against the first enemy. Moreover, we illustrate that the global defence against one enemy can be lower when only this enemy is present: this has important implications for experimental measures of resistance, and for organisms that invade an area with less enemies or whose community of enemies is reduced. We discuss possible implications of the existence of multiple enemies for conservation biology, biological control and chemical control.     

Natural variation in the contribution of microbial density to inducible immune dynamics

Immune responses evolve to balance the benefits of microbial killing against the costs of autoimmunity and energetic resource use. Models that explore the evolution of optimal immune responses generally include a term for constitutive immunity, or the level of immunological investment prior to microbial exposure, and for inducible immunity, or investment in immune function after microbial challenge. However, studies rarely consider the functional form of inducible immune responses with respect to microbial density, despite the theoretical dependence of immune system evolution on microbe‐ versus immune‐mediated damage to the host. In this study, we analyse antimicrobial peptide (AMP) gene expression from seven wild‐caught flour beetle populations (Tribolium spp.) during acute infection with the virulent bacteria Bacillus thuringiensis (Bt) and Photorhabdus luminescens (P.lum) to demonstrate that inducible immune responses mediated by the humoral IMD pathway exhibit natural variation in both microbe density‐dependent and independent temporal dynamics. Beetle populations that exhibited greater AMP expression sensitivity to Bt density were also more likely to die from infection, while populations that exhibited higher microbe density‐independent AMP expression were more likely to survive P. luminescens infection. Reduction in pathway signalling efficiency through RNAi‐mediated knockdown of the imd gene reduced the magnitude of both microbe‐independent and dependent responses and reduced host resistance to Bt growth, but had no net effect on host survival. This study provides a framework for understanding natural variation in the flexibility of investment in inducible immune responses and should inform theory on the contribution of nonequilibrium host‐microbe dynamics to immune system evolution.     

Manipulation of the host cell death pathway by Shigella

Host cells deploy multiple defences against microbial infection. One prominent host defence mechanism, the death of infected cells, plays a pivotal role in clearing damaged cells, eliminating pathogens, removing replicative niches, exposing intracellular bacterial pathogens to extracellular immune surveillance and presenting bacteria‐derived antigens to the adaptive immune system. Although cell death can occur under either physiological or pathophysiological conditions, it acts as an innate defence mechanism against bacterial pathogens by limiting their persistent colonization. However, many bacterial pathogens, including Shigella, have evolved mechanisms that manipulate host cell death for their own benefit.     

Do Induced Responses Mediate the Ecological Interactions Between the Specialist Herbivores and Phytopathogens of an Alpine Plant?

Plants are not passive victims of the myriad attackers that rely on them for nutrition. They have a suite of physical and chemical defences, and are even able to take advantage of the enemies of their enemies. These strategies are often only deployed upon attack, so may lead to indirect interactions between herbivores and phytopathogens. In this study we test for induced responses in wild populations of an alpine plant (Adenostyles alliariae) that possesses constitutive chemical defence (pyrrolizidine alkaloids) and specialist natural enemies (two species of leaf beetle, Oreina elongata and Oreina cacaliae, and the phytopathogenic rust Uromyces cacaliae). Plants were induced in the field using chemical elicitors of the jasmonic acid (JA) and salicylic acid (SA) pathways and monitored for one month under natural conditions. There was evidence for induced resistance, with lower probability and later incidence of attack by beetles in JA-induced plants and of rust infection in SA-induced plants. We also demonstrate ecological cross-effects, with reduced fungal attack following JA-induction, and a cost of SA-induction arising from increased beetle attack. As a result, there is the potential for negative indirect effects of the beetles on the rust, while in the field the positive indirect effect of the rust on the beetles appears to be over-ridden by direct effects on plant nutritional quality. Such interactions resulting from induced susceptibility and resistance must be considered if we are to exploit plant defences for crop protection using hormone elicitors or constitutive expression. More generally, the fact that induced defences are even found in species that possess constitutively-expressed chemical defence suggests that they may be ubiquitous in higher plants.     

The within‐host dynamics of infection in trans‐generationally primed flour beetles

Many taxa exhibit plastic immune responses initiated after primary microbial exposure that provide increased protection against disease‐induced mortality and the fitness costs of infection. In several arthropod species, this protection can even be passed from parents to offspring through a phenomenon called trans‐generational immune priming. Here, we first demonstrate that trans‐generational priming is a repeatable phenomenon in flour beetles (Tribolium castaneum) primed and infected with Bacillus thuringiensis (Bt). We then quantify the within‐host dynamics of microbes and host physiological responses in infected offspring from primed and unprimed mothers by monitoring bacterial density and using mRNA‐seq to profile host gene expression, respectively, over the acute infection period. We find that priming increases inducible resistance against Bt around a critical temporal juncture where host septicaemic trajectories, and consequently survival, may be determined in unprimed individuals. Our results identify a highly differentially expressed biomarker of priming, containing an EIF4‐e domain, in uninfected individuals, as well as several other candidate genes. Moreover, the induction and decay dynamics of gene expression over time suggest a metabolic shift in primed individuals. The identified bacterial and gene expression dynamics are likely to influence patterns of bacterial fitness and disease transmission in natural populations.     

Ontogenetic change in effectiveness of chemical defence against different predators in Oxycarenus true bugs

Many prey species change their antipredator defence during ontogeny, which may be connected to different potential predators over the life cycle of the prey. To test this hypothesis, we compared reactions of two predator taxa – spiders and birds – to larvae and adults of two invasive true bug species, Oxycarenus hyalinipennis and Oxycarenus lavaterae (Heteroptera: Oxycarenidae) with life-stage-specific chemical defence mechanisms. The reactions to larvae and adults of both true bug species strikingly differed between the two predator taxa. The spiders were deterred by the defences of adult bugs, but the larval defences were ineffective against them. By contrast, birds attacked the larvae considerably less often than the adult bugs. The results indicate a predator-specific ontogenetic change in defence effectiveness of both Oxycarenus species. The change in defence is likely linked to the life-stage-specific composition of secretions in both species: whereas secretions of larvae are dominated by unsaturated aldehydes, secretions of adults are rich in terpenoids, which probably serve dual function of defensive chemicals and pheromones. Our results highlight the variation in defence between different life stages and the importance of testing responses of different types of predators.     

Staphylococcus aureus protects its immune‐evasion proteins against degradation by neutrophil serine proteases

Neutrophils store large quantities of neutrophil serine proteases (NSPs) that contribute, via multiple mechanisms, to antibacterial immune defences. Even though neutrophils are indispensable in fighting Staphylococcus aureus infections, the importance of NSPs in anti‐staphylococcal defence is yet unknown. However, the fact that S. aureus produces three highly specific inhibitors for NSPs [the extracellular adherence proteins (EAPs) Eap, EapH1 and EapH2], suggests that these proteases are important for host defences against this bacterium. In this study we demonstrate that NSPs can inactivate secreted virulence factors of S. aureus and that EAP proteins function to prevent this degradation. Specifically, we find that a large group of S. aureus immune‐evasion proteins is vulnerable to proteolytic inactivation by NSPs. In most cases, NSP cleavage leads to functional inactivation of virulence proteins. Interestingly, proteins with similar immune‐escape functions appeared to have differential cleavage sensitivity towards NSPs. Using targeted mutagenesis and complementation analyses in S. aureus, we demonstrate that all EAP proteins can protect other virulence factors from NSP degradation in complex bacterial supernatants. These findings show that NSPs inactivate S. aureus virulence factors. Moreover, the protection by EAP proteins can explain why this antibacterial function of NSPs was masked in previous studies. Furthermore, our results indicate that therapeutic inactivation of EAP proteins can help to restore the natural host immune defences against S. aureus.     

Plasticity in the expression of direct and indirect defence traits of young plants of <Emphasis Type="Italic">Mallotus japonicus</Emphasis> in relation to soil nutritional conditions

Although soil nutrients can influence the defence strategy of plants that have multiple defence traits, to date, there have been few studies to examine this. To evaluate the effect of soil nutrients on multiple plant defences, we cultivated Mallotus japonicus under three soil nutritional conditions in the field, and experimentally examined the expression of a physical defence trait (trichomes), chemical traits (pellucid dots), and biotic traits (extrafloral nectaries (EFNs) and pearl bodies) of the plants, and the number of ants visiting them. Under the low soil nutritional condition, plants strongly expressed the physical defence by trichomes and chemical defence by pellucid dots, meaning that the young plants adopted direct defences under the poor soil nutritional condition. Under the high soil nutritional condition, in contrast, the plants strongly expressed the indirect defence traits. They produced abundant EFNs and pearl bodies, and attracted many ants. These results suggest that young plants of M. japonicus use different defence modes in response to different soil nutritional conditions.     

Herbivores and plant defences affect selection on plant reproductive traits more strongly than pollinators

Pollinators and herbivores can both affect the evolutionary diversification of plant reproductive traits. However, plant defences frequently alter antagonistic and mutualistic interactions, and therefore, variation in plant defences may alter patterns of herbivore‐ and pollinator‐mediated selection on plant traits. We tested this hypothesis by conducting a common garden field experiment using 50 clonal genotypes of white clover (Trifolium repens) that varied in a Mendelian‐inherited chemical antiherbivore defence—the production of hydrogen cyanide (HCN). To evaluate whether plant defences alter herbivore‐ and/or pollinator‐mediated selection, we factorially crossed chemical defence (25 cyanogenic and 25 acyanogenic genotypes), herbivore damage (herbivore suppression) and pollination (hand pollination). We found that herbivores weakened selection for increased inflorescence production, suggesting that large displays are costly in the presence of herbivores. In addition, herbivores weakened selection on flower size but only among acyanogenic plants, suggesting that plant defences reduce the strength of herbivore‐mediated selection. Pollinators did not independently affect selection on any trait, although pollinators weakened selection for later flowering among cyanogenic plants. Overall, cyanogenic plant defences consistently increased the strength of positive directional selection on reproductive traits. Herbivores and pollinators both strengthened and weakened the strength of selection on reproductive traits, although herbivores imposed ~2.7× stronger selection than pollinators across all traits. Contrary to the view that pollinators are the most important agents of selection on reproductive traits, our data show that selection on reproductive traits is driven primarily by variation in herbivory and plant defences in this system.     

Natural selection on quantitative immune defence traits: a comparison between theory and data

Parasites present a threat for free‐living species and affect several ecological and evolutionary processes. Immune defence is the main physiological barrier against infections, and understanding its evolution is central for predicting disease dynamics. I review theoretical predictions and empirical data on natural selection on quantitative immune defence traits in the wild. Evolutionary theory predicts immune traits to be under stabilizing selection owing to trade‐offs between immune function and life‐history traits. Empirical data, however, support mainly positive directional selection, but also show variation in the form of selection among study systems, immune traits and fitness components. I argue that the differences between theory and empirical data may at least partly arise from methodological difficulties in testing stabilizing selection as well as measuring fitness. I also argue that the commonness of positive directional selection and the variation in selection may be caused by several biological factors. First, selection on immune function may show spatial and temporal variation as epidemics are often local/seasonal. Second, factors affecting the range of phenotypic variation in immune traits could alter potential for selection. Third, different parasites may impose different selective pressures depending on their characteristics. Fourth, condition dependence of immune defence can obscure trade‐offs related to it, thus possibly modifying observed selection gradients. Fifth, nonimmunological defences could affect the form of selection by reducing the benefits of strong immune function. To comprehensively understand the evolution of immune defence, the role of above factors should be considered in future studies.