Inter- and intra-sexual variation in immune defence in the cabbage white butterfly, Pieris rapae L. (Lepidoptera: Pieridae)

Abstract.  1. Immune defence imposes fitness costs as well as benefits, so organisms are expected to optimise, not maximise, their immune responses. This should result in variation in immune responses under varying environmental conditions.
2. Males and females are expected to exhibit different immune responses because life-history differences between the sexes affect optimal immune response. These life-history differences should usually result in a greater female, than male, immune defence. In this study, intra- and inter-sexual variation in one component of immune defence, the encapsulation response, in cabbage white butterflies ( Pieris rapae L.), was examined.
3. Encapsulation decreased with increasing age and in response to reduced diet quality.
4. Contrary to predictions, males generally had greater immune responses than females, although this pattern varied with age.
5. These patterns of inter- and intra-sexual variation in encapsulation may result from resource-based trade-offs with components of reproductive effort and/or because of sexual dimorphism in melanin-based wing patterns.     

Examination of the immune responses of males and workers of the leaf-cutting ant Acromyrmex echinatior and the effect of infection

Summary. Parasites represent significant challenges to social insects. The high density, interaction rate and relatedness of individuals within colonies are all predicted to make social insect colonies particularly vulnerable to parasites. To cope with this pressure, social insects have evolved a number of defence mechanisms. These include the immune response, which, aside from in bumblebees, has been relatively little studied in social insects. Here we compare the immune responses of males and workers of the leaf-cutting ant Acromyrmex echinatior and examine the effect upon immunocompetence of prior exposure to a virulent parasite. Males have a far lower immune response than workers, suggesting either haploid susceptibility or reduced investment in immunity by the short-lived males. There was also significantly less variation in the immune response of males than of workers, which may be due to leaf-cutting ant workers being more variable in age or more genetically diverse within colonies. When exposed to the entomopathogenic fungus Metarhizium, workers expressed a substantially reduced immune response 96 h after infection, suggesting that the immune system was either depleted by having to respond to the Metarhizium infection or was depressed by the parasite. The results suggest that the immune response is a costly and limited process, but further experiments are needed to distinguish between the alternative explanations for the effects observed.Received 3 August 2004; revised 3 February 2005; accepted 2 March 2005.     

A trade-off between sexual signalling and immune function in a natural population of the drumming wolf spider Hygrolycosa rubrofasciata

The field of ecological immunology is ultimately seeking to address the question ‘Why is there variation in immune function?’ Here, we provide experimental evidence that costs of ubiquitous sexual signals are a significant source of variation in immune function. In the mating season, males of the wolf spider Hygrolycosa rubrofasciata drum against dry leaves while wandering around the habitat searching for receptive females. According to a previous study, the male metabolic rate during the drumming increases 22‐fold compared to the resting metabolic rate. In the present study, we examined whether investment in costly courtship drumming decreases male immune function in a wild population of H. rubrofasciata. We induced males to increase their drumming rate by introducing females in proximity. As estimates of male immune function, we used lytic activity and encapsulation rate. Lytic activity estimates the concentration of antimicrobial peptides in haemolymph, which have been shown to play an important role in defence against bacteria, viruses and fungi. Encapsulation is an important defence mechanism against nematodes and insect parasitoids, but it also plays a role in defence against viruses. Our results show that males with nonarbitrarily increased investment in drumming rate had considerably lower lytic activities than control males. Also, there was a tendency for males with nonarbitrarily increased investment in drumming rate to have lower encapsulation rates than control males. This study provides experimental evidence for the first time, to our knowledge, that there are direct immunological costs of sexual signalling in natural populations. Therefore, immunological costs of sexual signals may provide significant phenotypic variation to parasite‐mediated sexual selection.     

Greater cuticular melanism is not associated with greater immunogenic response in adults of the polymorphic mountain stone weta, Hemideina maori

Abstract.  1. Greater immune function is associated with the high-density melanic phase of polyphenic insects, appearing to compensate for density-dependent increases in susceptibility to parasites and/or pathogens. Other types of discrete variation in cuticular colour occur in insects (which may or may not be associated with melanin pigmentation), but whether this variation is predictive of immune ability has not been investigated.
2. In the mountain stone weta Hemideina maori , a black morph and yellow banded morph occur. These morphs are not seasonally polyphenic and have discrete haplotype genetic markers. Black individuals are typically found at lower local densities than yellow individuals, contrary to relations between cuticular melanism and density seen in polyphenic insects.
3. Yellow males and females had greater melanotic encapsulation responses upon immune challenge than did black males and females, but these differences were not associated with differences in temperature selection between morphs. Morph differences in melanotic encapsulation responses were somewhat related to differences between morphs in haemocyte concentrations.
4. These results indicate that a common form of immune expression is not heightened with dark coloration in the mountain stone weta. Thus, earlier findings of greater immunity associated with darker cuticles in phase polyphenic insects cannot be extended to insects with other forms of discrete colour variation. These findings will help in elucidating causes and consequences of such colour polymorphism, which is widespread in several insect orders.     

Inbred and outbred honey bees (Apis mellifera) have similar innate immune responses

Social insect colonies provide ideal conditions for the spread of pathogens. It has been proposed that the extreme polyandry and genetic diversity seen in the colonies of some eusocial insect species is central to a colony’s defence against disease. Indeed, empirically, colonies headed by polyandrous queens have lower incidence of pathogens than genetically uniform monoandrous colonies. The mechanisms of improved resistance in genetically diverse colonies could arise from the genetic diversity among worker genotypes or from increased innate immunity arising from heterozygosity at immune gene loci within individual workers. Here, we investigate the effects of heterozygosity on two components of the honey bee (Apis mellifera) innate immune system: encapsulation and phenoloxidase (PO) activity. No significant effect of heterozygosity on immune system activity was evident for either encapsulation or PO activity. Thus, we conclude that while encapsulation and PO activity are important components of the immune response, it seems that they do not underlie the positive effects of genetic diversity on parasite and pathogen resistance in honey bees.     

Bottom-up cascading effects in a tritrophic system: interactions between plant quality and host-parasitoid immune responses

Abstract.  1. Little is known about underlying mechanisms by which plants indirectly affect parasitism success in hymenopteran endoparasitoids. The hypothesis that host-plant effects can challenge the innate immune system of an insect host was experimentally tested in this study using a model tritrophic, crucifer – lepidopteran [ Plutella xylostella (L.)] – parasitoid [ Cotesia plutellae (Kurdjumov)], system.
2. The effects of host-plant suitability on herbivore performance and parasitism were examined. The bottom-up effect of plant suitability on host-parasitoid immune responses was then evaluated using measures of cellular and humoral effectors.
3. Host-plant quality showed a significant effect on the encapsulation response of P. xylostella to first instar but not to second instar parasitoid larvae. Encapsulation was never sufficient to prevent parasitoid emergence.
4. Poor host-plant suitability suppressed phenoloxidase activity in the absence of the parasitoid. The suppressive effect of C. plutellae on phenoloxidase activity was much greater and no plant effects were detectable after insects had been parasitized.
5. Despite strong plant effects on parasitism, those on immune effectors of the host were transitory or overwhelmed by the effect of the parasitoid.
6. These results demonstrated that plant-mediated variation in parasitism success by C. plutellae were not as a result of plant nutritional status or other attributes affecting the immune function of P. xylostella , nor to host-plant effects on superparasitism.
7. In these experiments, P. xylostella was a fully permissive host to C. plutellae and host-plant-mediated effects on the innate immune response appeared to play no part in parasitoid survival within hosts.     

Senescence of immune defence in Bombus workers

Abstract 1. Senescence in workers of social insects is a particularly intriguing life-history trait as the future fitness of workers relies primarily on age-dependent survival rate. The pattern of senescence of immune defence traits was investigated under laboratory conditions in workers of two bumble bees: Bombus terrestris and B. lucorum .
2. In both species, there was a significant decrease with age in the ability to encapsulate a foreign object (a global measure of the efficiency of immune systems). This pattern of senescence was observed in all colonies in B. terrestris (seven) and B. lucorum (eight) assayed, even though, for the latter, there was some heterogeneity among colonies.
3. In B. terrestris , two other measures of immune defence were taken: the relative percentage of fat body in the abdomen and the concentration of haemocytes (the immune defence cells). The quantity of fat body increased only slightly with age and there was no effect for the concentration of haemocytes. Interestingly, the concentration of haemocytes decreased strongly after an encapsulation response, regardless of the age of workers.
4. The importance of the senescence pattern observed for the immune defence traits is discussed in the context of the social biology of workers.     

Rapid induction of immune density-dependent prophylaxis in adult social insects

The innate immune system provides defence against parasites and pathogens. This defence comes at a cost, suggesting that immune function should exhibit plasticity in response to variation in environmental threats. Density-dependent prophylaxis (DDP) has been demonstrated mostly in phase-polyphenic insects, where larval group size determines levels of immune function in either adults or later larval instars. Social insects exhibit extreme sociality, but DDP has been suggested to be absent from these ecologically dominant taxa. Here we show that adult bumble-bee workers (Bombus terrestris) exhibit rapid plasticity in their immune function in response to social context. These results suggest that DDP does not depend upon larval conditions, and is likely to be a widespread and labile response to rapidly changing conditions in adult insect populations. This has obvious ramifications for experimental analysis of immune function in insects, and serious implications for our understanding of the epidemiology and impact of pathogens and parasites in spatially structured adult insect populations.     

Immunity and reproduction during colony foundation in the dampwood termite, Zootermopsis angusticollis

Abstract.  Termite primary reproductives may be exposed to pathogens when dispersing from their parental nest and establishing a new colony. Immunity and reproduction are investigated during colony foundation by implanting a nylon filament into the abdomen of mated and unmated female and male primary reproductives of the dampwood termite Zootermopsis angusticollis. Primary reproductives are paired in combinations of female/male, female/female and male/male and, using confocal microscopy, immune defence is assessed by measuring the degree of encapsulation of nylon implants during three periods of colony foundation: (I) shortly after pairing; (II) during copulation/oocyte maturation; and (III) during oviposition. There are differences in the encapsulation response of mated and unmated termites that are contingent on the period of colony foundation when termites are challenged. Mated females and males have significantly greater encapsulation responses than their unmated counterparts shortly after pairing, perhaps as a prophylactic measure against exposure to disease. The encapsulation response of mated and unmated males does not differ significantly during periods II and III. The onset of oviposition is significantly delayed in mated females that received implants during periods I and II. Mated females have a significantly reduced encapsulation response during the time of copulation and oocyte maturation, but not during oviposition. Overall, males have a significantly greater ability than females to encapsulate a nylon implant. The findings suggest that reproduction can reduce the immune response in female primary reproductives. The results are discussed in light of trade-offs between immunity and reproduction during the critical life-history phase of colony establishment in termites.     

Colony performance and immunocompetence of a social insect, Bombus terrestris, in poor and variable environments

1. Feeding and defence against parasites are among the major simultaneous requirements for survival and successful reproduction of any animal, including colonies of social insects. Harsh environments may therefore not only negatively affect overall success, but force animals to allocate resources in different ways, for example, to reduce immune defence under adverse conditions while maintaining growth rate.
2. This study examines how colony growth, reproduction and immune defence of the bumble bee, Bombus terrestris , varies with environmental condition. In particular, the manner in which limited or unpredictable food availability affects the number and size of workers and sexuals produced, the timing of reproduction, and the level of encapsulation of a novel antigen was tested. A new experimental paradigm was applied by splitting colonies into two halves while ensuring integrity by regular swaps of the residing queen.
3. The results show that limited food availability, as expected, led to fewer and smaller workers and sexuals. Unpredictable food availability, in contrast, was associated with higher numbers of workers and heavier sexuals. Also, reproductive success was highest under unpredictable conditions, due to an increased rate of collection of resources.
4. The timing of reproduction did not vary among treatments. Contrary to expectation, also immunocompetence did not vary among treatments.     

Inbreeding and extreme outbreeding cause sex differences in immune defence and life history traits in Epirrita autumnata

Empirical studies in vertebrates support the hypothesis that inbreeding reduces resistance against parasites and pathogens. However, studies in insects have not found any evidence that inbreeding compromises immune defence. Here we tested whether one generation of brother-sister mating or extreme outbreeding (mating between two populations) have an effect on innate immunity and life history traits in the autumnal moth, Epirrita autumnata. We show that the effect of inbreeding on immune response differed between the sexes: whereas in females, inbreeding significantly reduced encapsulation response against nylon monofilament ability, it did not have a significant effect on male immune response. There were also differences in the correlation of the immune response with other traits: in females increased immune response was positively correlated with large size, whereas in males immune response increased with a reduction in development time. Immune response differed significantly among families in males but not in females, both for the inbreeding and extreme outbreeding experiments. In conjunction with the observed immune responses to inbreeding, these data suggest that in males genetic variation for immune response is largely additive or non-directional with respect to dominance, whereas in females variation is much reduced and consists of directional dominance variance. Further, we show that encapsulation response against nylon monofilament is associated with the resistance against real pathogens suggesting that this widely used method to measure the strength of immune defence in insects is also a biologically relevant method.     

On understanding seasonal increases in damselfly defence and resistance against ectoparasitic mites

Abstract.  1. Defence against parasites and pathogens can be essential, yet not all hosts respond similarly to parasitic challenge. Environmental conditions are thought to explain variation in host responses to parasites.
2.  Lestes forcipatus damselflies emerging later in the season have shown higher resistance to the mite, Arrenurus planus , than hosts emerging earlier. This study was undertaken to determine whether variation in environmental temperatures characteristic of early vs. late emergence times, degree or costs of mite parasitism, and/or size of newly emerged adults could explain seasonal variation in defence and resistance to ectoparasitic mites.
3. In this study damselflies from early vs. late emergence groups differed in size at emergence and mite intensity. In general, early hosts were larger and had more mites than later hosts. However only experimental temperatures experienced by damselflies at emergence influenced defence and resistance against mites and not host size or degree of parasitism.
4. More specifically, hosts from early and late emergence groups did not differ in defence and resistance when held at the same temperatures in incubators. Housing at a high temperature, indicative of later in the season, was associated with higher defence and resistance for damselflies from both early and late emergence groups.
5. These results indicate that daily temperatures in relation to emergence timing can account for seasonal increases in resistance for this temperate insect. Seasonal increases in resistance may be expected for other temperate insect–parasite associations and should have important implications for the phenology of parasites and for seasonal variation in parasite-mediated selection.     

Effects of selective episodes in the field on life history traits in the bumblebee Bombus terrestris

Natural selection has different fitness consequences when it acts during different life cycle stages of an organism. In social insects, for example, the colony foundation and early colony growth is a critical time period with high probability of failure. Here we used colonies of the bumblebee Bombus terrestris L. to test whether selective episodes at different life cycle stages result in differences in colony performance and fitness. The timing of a selective episode was varied by field exposure of colonies, either permanently or during a short period at three different colony life cycle stages – early, middle, or late in the cycle. We found that selective episodes at different life cycle stages did not affect maximal size, fitness or survival of colonies, or the time span between colony foundation and reproduction. Instead, the colonies were able to compensate for costs encountered by delaying reproduction. This might have important fitness consequences, since later emerging sexuals might be faced with harsher environmental conditions and increased costs of finding a mate. In addition, an important component of selection might be parasitism and the resulting resource allocation to the immune system. We here measured the generalized immune response (i.e. encapsulation response) of early produced workers as an indicator of a colony's capacity to defend against parasitism. Encapsulation response correlated positively with eventual colony size and fitness, indicating that this measure of "immunocompetence" correlates with important life history traits.     

Temporal dynamics of the encapsulation response towards a synthetic immune challenge in Acheta domesticus

Abstract.  Researchers interested in insect 'ecological immunology' often quantify variation among individuals in the capacity to produce an immune response by challenging the immune system with a synthetic implant. Many studies focus on the encapsulation response in particular, because it requires the coordination of different cellular and humoral factors. However, most encapsulation assays are based on a single assay period, in part because the assay is destructive, such that the response cannot be measured repeatedly in the same individual. This approach may be problematic. For example, if the time-course of the response is complex, an arbitrarily chosen assay period may fail to capture the most biologically relevant parameters of the response. In the present study, the time-course of the encapsulation response towards a synthetic, nylon implant is investigated over a 5-day period, using Acheta domesticus as a model host. Encapsulation is quantified at intervals by randomly allocating crickets to different time-points. The most rapid period of capsule development appears to occur during the first 6-h period. Subsequent increases in capsule size can only be confirmed during the latter stages of the assay period, when there is also a significant increase in haemocyte density. The results indicate that, in any given system, the assay period should ideally be based on preliminary data quantifying the time-course of the response. Such data encourage clearer conclusions regarding the biological meaning of the assay and will increase relevance for real-host parasite systems, in which temporal dynamics can be highly variable.     

Covariation between colony social structure and immune defences of workers in the ant Formica selysi

Several ant species vary in the number of queens per colony, yet the causes and consequences of this variation remain poorly understood. In previous experiments, we found that Formica selysi workers originating from multiple-queen (=polygyne) colonies had a lower resistance to a fungal pathogen than workers originating from single-queen (=monogyne) colonies. In contrast, group diversity improved disease resistance in experimental colonies. This discrepancy between field and experimental colonies suggested that variation in social structure in the field had antagonistic effects on worker resistance, possibly through a down-regulation of the immune system balancing the positive effect of genetic diversity. Here, we examined if workers originating from field colonies with alternative social structure differed in three major components of their immune system. We found that workers from polygyne colonies had a lower bacterial growth inhibitory activity than workers from monogyne colonies. In contrast, workers from the two types of colonies did not differ significantly in bacterial cell wall lytic activity and prophenoloxidase activity. Overall, the presence of multiple queens in a colony correlated with a slight reduction in one inducible component of the immune system of individual workers. This reduced level of immune defence might explain the lower resistance of workers originating from polygyne colonies despite the positive effect of genetic diversity. More generally, these results indicate that social changes at the group level can modulate individual immune defences.     

Social immunity

Social insect colonies have evolved collective immune defences against parasites. These 'social immune systems' result from the cooperation of the individual group members to combat the increased risk of disease transmission that arises from sociality and group living. In this review we illustrate the pathways that parasites can take to infect a social insect colony and use these pathways as a framework to predict colony defence mechanisms and present the existing evidence. We find that the collective defences can be both prophylactic and activated on demand and consist of behavioural, physiological and organisational adaptations of the colony that prevent parasite entrance, establishment and spread. We discuss the regulation of collective immunity, which requires complex integration of information about both the parasites and the internal status of the insect colony. Our review concludes with an examination of the evolution of social immunity, which is based on the consequences of selection at both the individual and the colony level.     

Can intra‐specific genetic variation in arbuscular mycorrhizal fungi (Glomus etunicatum) affect a mesophyll‐feeding herbivore (Tupiocoris notatus Distant)?

Abstract.  1. Arbuscular mycorrhizal fungal (AMF) infection can have negative, positive or neutral effects on insect herbivore populations, but patterns are difficult to predict.
2. Intra-specific genetic variation in nutrient uptake ability between fungal isolates may also have indirect effects on insect herbivores due to changes in plant quality. In preliminary studies mirid ( Tupiocoris notatus ) populations were significantly reduced on tobacco ( Nicotiana rustica ) colonised by AMF but it was unknown if same-species fungal isolates differed in their effect.
3. An experiment was performed as a first test of the effect of intra-specific genetic variation in the mycorrhizal fungus Glomus etunicatum on mirid nymphal population structure, dynamics, and growth rate.
4. Mirid nymphal populations were lower on mycorrhizal fungal-infected plants. Population size, however, did not differ between the mycorrhizal isolates. While no statistical difference in population between isolates was found, one isolate consistently had 1.7–2.4 times lower mirid populations compared with the controls, indicating that the magnitude of effect is different between mycorrhizal isolates.
5. The significantly negative effect of AMF on mirid populations likely resulted from AMF-induced changes in plant quality (e.g. increased defence). This study lends further support to recent demonstrations that below-ground symbionts significantly influence above-ground processes. In addition, mycorrhizal fungi can affect insect population structure, which may have consequences for future herbivory.     

Colony pollen reserves affect body size,sperm production and sexual development in males of the stingless bee <Emphasis Type="Italic">Melipona beecheii</Emphasis>

The production of male sexual offspring by social insect colonies is often strongly seasonal or resource-dependent. In stingless bees, males are produced in smaller numbers under conditions of low colony food reserves; whether such males are negatively affected in traits related to reproductive success is not known. We compared body size, sperm production and sexual maturity in Melipona beecheii males reared with experimentally supplemented or reduced pollen reserves, but with otherwise equal numbers of workers and equal quantities of honey reserves. We also studied the same traits in males collected from non-manipulated colonies with pollen reserves intermediate between the supplemented or reduced groups but with more workers and honey reserves. Males reared under experimentally reduced pollen reserves had significantly smaller bodies and lower sperm counts compared to those reared in colonies with experimentally supplemented pollen reserves. There was also a significantly positive relationship between the number of sperm and body size in males across all colony treatments. The maximum number of sperm in seminal vesicles was recorded 2 days later in males from colonies with reduced pollen compared to males from colonies with supplementary pollen. Males from non-manipulated colonies were intermediate in size, sperm count and speed of maturation. Our study documents for the first time the existence of large size variation in males of stingless bees that is related with the amount of pollen reserves in their natal colony. We conclude that a colony’s pollen reserves have a major impact on male body size, sperm production and speed of sexual maturity in this stingless bee, which may be the case in other social insects. Stingless bees are a good model system to study the balance between colony-level selection and individual-level selection on male sexually selected traits such as body size.     

Mating status, immune defence, and multi-parasite burden in the damselfly Coenagrion armatum

Immunity and reproductive effort are both physiologically costly and often a trade-off between these functions has been shown. In studies with damselflies, parasite load has been associated with fitness costs, such as reductions in mating success, male condition, and survival. Although each individual may be simultaneously infected by various parasite species, most studies have concentrated on the effects of a single parasite taxon. We examined natural ecto- and endoparasite infection levels in male Coenagrion armatum (Charpentier) (Odonata: Coenagrionidae) damselflies in relation to their mating status, fat reserves, and ability to further mount an immune response, measured as encapsulation of an experimentally introduced foreign object. Encapsulation response was lower for mated (paired) males than for single males and declined with increasing water mite abundance. Mated males had fewer water mites than single males. Male weight or fat reserves did not explain variation in encapsulation response. The number of gregarine gut parasites was not related to the level of encapsulation response and did not differ between mated and single males. However, there was a negative correlation between mite abundance and gregarine load. Our data suggest that current mite infection may compromise a male's resistance against further infections by pathogens and parasites, and there may be a trade-off between reproductive effort and encapsulation response in male C. armatum .     

Sexually dimorphic effect of mating on the melanotic encapsulation response in the harem‐defending Wellington tree weta,Hemideina crassidens (Orthoptera: Tettigonioidea: Anostostomatidae)

Reproductive activities are generally costly to immune responsiveness because limited resources required by reproduction are diverted away from immunity (and vice versa). Reproduction, however, is not expected to affect the immune response in males and females similarly as mating is expected to negatively affect male immunity more so than female immunity. Here, I test the phenotypic plasticity hypothesis in the Wellington tree weta (Hemideina crassidens), a sexually dimorphic orthopteran insect that is endemic to New Zealand. My laboratory experiment showed that although males had higher rates of melanotic encapsulation than females, contrary to prediction, females were the only sex significantly affected by mating and the effect was positive. In addition to immunity differing between the sexes, immune function can differ intrasexually, particularly when males are polymorphic and different investment strategies are used to maximize fitness. Male H. crassidens exhibit alternative mating strategies that are represented by three different morphotypes. I therefore explored whether the morphs differed in their melanotic encapsulation response and whether mating affected the morphs differently. I found no difference among morphs or an effect of mating on male immune response.