Ecological Immunology

The innate immune system in insects is composed of a large variety of specific and non-specific responses that are activated in response to the presence of foreign agents. One important element in such responses is the enzyme phenoloxidase (PO). Here, we review recent progress in PO research and discuss new applications in the emerging field of ecological immunology. Phenoloxidase produces indole groups, which are subsequently polymerized to melanin. The enzymatic reactions in turn produce a set of intermediate products such as quinones, diphenols, superoxide, hydrogen peroxide, and reactive nitrogen intermediates, which are important during defense against bacterial (gram+ and )), fungal, and viral agents. Phenoloxidase requires a complex system of activation and inhibition that involves various cell types, PO zymogens, inhibitor enzymes, and signaling molecules. Finally, research in evolutionary ecology has studied the costs of PO in terms of resource use and pleiotropic relations with other key traits and functions. These studies indicate that PO is a costly trait, whose production and maintenance have fitness costs for hosts. Phenoloxidase does not seem to be an indicator of resistance but rather of host condition. Finally, we put forward some basic directions for future investigation of PO aimed at explaining its activating system, its substrates, its coordination with other immune components to fight off pathogens, and variation in PO in relation to gender, life stages, seasonality, and across different host species.

Sexual selection theory indicates that males use sexual traits to signal immune ability, a hypothesis known as the immunocompetence principle. A positive relationship between sexual traits and immune ability is not always present. Here we illustrate this pattern by using five damselfly species in the genus Hetaerina. Previous studies have documented a positive correlation between sexual trait expression (wing spot size) and immune ability in members of this genus. These studies have also documented that there are fitness and energetic costs of producing and bearing wing pigmentation. First we used five Hetaerina species to investigate the correlation between spot size and phenoloxidase (PO) activity (a key insect immune component) in two contrasting seasons. Second, we experimentally challenged males of two Hetaerina species and correlated spot size with PO activity and melanization ability. Results indicate either a positive relationship, a negative relationship or, more commonly, no relationship at all between immune components and wing pigmentation. Season did not predict any of these relationships or expression of spot size and PO activity. These results, although limited to two immune components, indicate that the relationship between sexual trait expression and immunity is not always consistent.

Sick animals may change their feeding behaviour to compensate for infections. However, there is little information regarding whether infection affects (1) feeding behaviour of predators, (2) feeding behaviour using an experimental approach in the wild, (3) other costly behaviours and/or (4) physiological components of condition. We experimentally infected males of the predatory damselfly Hetaerina americana in a field experiment. We hypothesized that infection would reduce feeding behaviour. We further predicted a reduction in territorial activity, an increase in immune response (measured by the activity of phenoloxidase, PO) and a reduction of fat reserves and flight-associated muscle mass (two traits usually traded off with immune ability and territorial behaviour). We also infected males in a laboratory experiment that controlled for food supply and territorial activity, and measured the same physiological characters. Immune challenges in the field experiment unexpectedly increased feeding rate but did not change territorial activities. Muscle mass was reduced in the field but not in the laboratory, probably because of differences in the presence of energetically expensive territorial activities. In the laboratory, starvation and infection reduced PO activity and fat stores but did not affect muscle mass. Thus, our field and laboratory results support the idea that increased feeding compensates for infections in predators. Ó

Avian migration has been shown to be a life
history strategy for surviving environmental resource variability,
but it requires increased body reserves for long
distance flight. Fat reserves make excellent energy stores
for barrier crossing, whereas proteins generate less energy
for the same mass of fat but provide water during breakdown,
which may become especially useful when birds
become water stressed. Intra-African migrants are probably
unlikely to have to cross barriers equivalent to the Sahara
and the Mediterranean sea and so may have different patterns
of mass reserves reflecting the utility of metabolizing
fat versus protein in hot, tropical environments. We examined
differences in proportions of body mass gain, pectoral
muscle score, and fat score between intra-African migrants,
Palearctic migrants, and resident African species. We tested
whether intra-African migrants show a distinct seasonal
peak in mass gain corresponding to expected peak migration
period in a manner similar to Palearctic migrants, but
maintain larger muscle tissues, because Palearctic migrants
are more constrained by a need to heavily up-regulate fat in
addition to fat-free reserves before migration due to the
energy requirements of crossing the barrier of the Sahara.
We found that intra-African migrants had a peak seasonal
mass gain similar to Palearctics whereas African residents
did not, and that Palearctics increased fat reserves with
pectoral muscle reserves, so that they had much higher fat
scores for any given level of pectoral muscle compared to
intra-African migrants or resident species. Our results
suggest that barrier crossing leads to a distinct increase in
fat reserves rather than migration per se, and suggests that
intra-African migrants are more similar in their reserve
management to African residents. Mass gain devoid of
visible fat accumulation in intra-African migrants may,
therefore, suggest absence of barriers during migration.

Parasites infect hosts non-randomly as genotypes of hosts vary in susceptibility to the same genotypes of parasites, but this specificity may be modulated by environmental factors such as nutrition. Nutrition plays an important role for any physiological investment. As immune responses are costly, resource limitation should negatively affect immunity through trade-offs with other physiological requirements. Consequently, nutritional limitation should diminish immune capacity in general, but does it also dampen differences among hosts? We investigated the effect of short-term pollen deprivation on the immune responses of our model host Bombus terrestris when infected with the highly prevalent natural parasite Crithidia bombi. Bumblebees deprived of pollen, their protein source, show reduced immune responses to infection. They failed to upregulate a number of genes, including antimicrobial peptides, in response to infection. In particular, they also showed less specific immune expression patterns across individuals and colonies. These findings provide evidence for how immune responses on the individual-level vary with important elements of the environment and illustrate how nutrition can functionally alter not only general resistance, but also alter the pattern of specific host-parasite interactions.

Sick animals may change their feeding behaviour to compensate for infections. However, there is little information regarding whether infection affects (1) feeding behaviour of predators, (2) feeding behaviour using an experimental approach in the wild, (3) other costly behaviours and/or (4) physiological components of condition. We experimentally infected males of the predatory damselfly Hetaerina americana in a field experiment. We hypothesized that infection would reduce feeding behaviour. We further predicted a reduction in territorial activity, an increase in immune response (measured by the activity of phenoloxidase, PO) and a reduction of fat reserves and flight-associated muscle mass (two traits usually traded off with immune ability and territorial behaviour). We also infected males in a laboratory experiment that controlled for food supply and territorial activity, and measured the same physiological characters. Immune challenges in the field experiment unexpectedly increased feeding rate but did not change territorial activities. Muscle mass was reduced in the field but not in the laboratory, probably because of differences in the presence of energetically expensive territorial activities. In the laboratory, starvation and infection reduced PO activity and fat stores but did not affect muscle mass. Thus, our field and laboratory results support the idea that increased feeding compensates for infections in predators. Ó

Migration is a physiologically demanding activity. Recent studies suggest that migrating birds can improve their immune and antioxidant status during stopover, implying that variation in stopover habitat can affect migrants' health. We studied 2 species that are strongly frugivorous during fall migration, Swainson's Thrushes (Catharus ustulatus) and Gray Catbirds (Dumetella carolinensis). We asked whether birds that used shrubland dominated by exotic plants experienced differences in mass change, immune function, and antioxidant status relative to conspecifics in native-dominated shrubland during fall stopover in Michigan, USA, in 2012–2013. We found no habitat-related differences in any of the measured health parameters for Swainson's Thrushes. However, Gray Catbirds using native-dominated shrubland retained mass while those in exotic-dominated shrubland lost mass in 2013. Gray Catbirds in exotic-dominated habitat that year also had poorer immune status (elevated granulocyte:lymphocyte ratio and reduced hemagglutination and haptoglobin) and had lower plasma total carotenoids (immunostimulatory antioxidants) relative to conspecifics in native-dominated shrubland. While these findings were not replicated in 2012, circulating antioxidant capacity was lower in both years in Gray Catbirds captured in exotic habitat. Habitat-specific estimates of dietary energy, carotenoid, and antioxidant content per unit mass of fruit were similar between habitat types for Swainson's Thrushes. For Gray Catbirds, however, the fruit diet was lower in carotenoids and antioxidant capacity, but not in energy, in exotic habitat. Our results provide evidence that differential use of stopover habitats may affect the immune and antioxidant status of migrating landbirds. Furthermore, our results suggest that habitat may affect health status based on the complex relationships among quality and abundance of food, food preferences, and refueling performance, which vary among species. We suggest that future studies measure immune and antioxidant metrics in addition to refueling performance to better understand the effects of stopover habitat use on landbird migrant health. This information may improve assessment of habitat quality for migrants.

Objectives: Despite public health concerns about hookworm infection in pregnancy , little is known about immune profiles associated with hookworm (Necator americanus and Ancylostoma duodenale) infection during pregnancy. Fetal tolerance requirements may constrain maternal immune response to hookworm, thereby increasing susceptibility to new infections or increasing hemoglobin loss. To explore this possibility, we study systemic immune response and hemoglobin levels in a natural fertility population with endemic helminthic infection. Methods: We used Bayesian multilevel models to analyze mixed longitudinal data on hemoglobin, hookworm infection, reproductive state, eosinophils, and erythro-cyte sedimentation rate (ESR) to examine the effects of pregnancy and hookworm infection on nonspecific inflammation, cellular parasite response, and hemoglobin among 612 Tsimane women aged 15-45 (1016 observations). Results: Pregnancy is associated with lower eosinophil counts and lower eosino-phil response to hookworm, particularly during the second and third trimesters. Both hookworm and pregnancy are associated with higher ESR, with evidence for an interaction between the two causing further increases in the first trimester. Pregnancy is moderately associated with higher odds of hookworm infection (OR: 1.23, 95% CI: 0.83 to 1.83). Pregnancy and hookworm both decrease hemoglobin and may interact to accentuate this effect in the first-trimester of pregnancy (Interaction: β: −0.30 g/dL; CI: −0.870 to 0.24). Conclusions: Our findings are consistent with a possible trade-off between hook-worm immunity and successful pregnancy, and with the suggestion that hookworm and pregnancy may have synergistic effects, particularly in the first trimester.

Parasites are characteristically aggregated within hosts, but identifying the mechanisms underlying such aggregation can be difficult in wildlife populations. We examined the influence of host age and sex over an annual cycle on the eggs per gram of feces (EPG) of nematode parasites infecting wild Japanese macaques (Macaca fuscata yakui) on Yakushima Island. Five species of nematode were recorded from 434 fecal samples collected from an age-structured group of 50 individually recognizable macaques. All parasites exhibited aggregated EPG distributions. The age-infection profiles of all three directly transmitted species (Oesophagostomum aculeatum, Strongyloides fuelleborni, and Trichuris trichiura) exhibited convex curves, but concavity better characterized the age-infection curves of the two trophically transmitted species (Streptopharagus pigmentatus and Gongylonema pulchrum). There was a male bias in EPG and prevalence of infection with directly transmitted species, except in the prevalence of O. aculeatum, and no sex bias in the other parasites. Infection with O. aculeatum showed a female bias in prevalence among young adults, and additional interactions with sex and seasonality show higher EPG values in males during the mating season (fall) but in females during the birth season (spring). These patterns suggest that an immunosuppressive role by reproductive hormones may be regulating direct, but not indirect, lifecycle parasites. Exposure at an early age may trigger an immune response that affects all nematodes, but trophically transmitted species appear to accumulate thereafter. Although it is difficult to discern clear mechanistic explanations for parasite distributions in wildlife populations, it is critical to begin examining these patterns in host species that are increasingly endangered by anthropogenic threats.

Immune responses are costly, causing trade-offs between defense and other host life history traits. Aphids present a special system to explore the costs associated with immune activation since they are missing several humoral and cellular mechanisms thought important for microbial resistance, and it is unknown whether they have alternative, novel immune responses to deal with microbial threat. Here we expose pea aphids to an array of heat-killed natural pathogens, which should stimulate immune responses without pathogen virulence, and measure changes in life-history traits. We find significant reduction in lifetime fecundity upon exposure to two fungal pathogens, but not to two bacterial pathogens. This finding complements recent genomic and immunological studies indicating that pea aphids are missing mechanisms important for bacterial resistance, which may have important implications for how aphids interact with their beneficial bacterial symbionts. In general, recent exploration of the immune systems of non-model invertebrates has called into question the generality of our current picture of insect immunity. Our data highlight that taking an ecological approach and measuring life-history traits to a broad array of pathogens provides valuable information that can complement traditional approaches.

Adaptive immunity allows vertebrates to gain protection against repeated pathogenic infections. Analogous responses (priming) have been recently uncovered in invertebrates. However, whether such responses are widespread is not known. The present study investigated the presence of immunological priming in males of a species whose phylogenetic position places it in one of the less derived insect orders. It is hypothesized that the efficiency of such a response could be related to animal condition, as assessed by the expression of a sexually selected ornament. Hetaerina americana Fabricius (Odonata: Calopterygidae) males bear a conspicuous ornament (a red wing spot), which is evolutionarily maintained via male territorial competition. Using field-collected animals, a group of males is challenged with bacteria before exposure to a higher dose of the same or a different bacteria, and survival is compared with that of infected males not previously challenged, as well as control groups. Gram-positive and Gram-negative bacteria are used. To explore how long priming may take to work, the second exposure is carried out either after 1 or 5 days. Red spot and body size are entered in the analysis as predictors of survival within and between groups. There is no difference in survival among groups, which suggests no priming effect. Overall, red spot and body size are not consistent in explaining survival.

Ecological and social factors are central in the emergence and transmission of infectious diseases, thus bearing the potential for shaping a species' immune functions. Although previous studies demonstrated a link between social factors and the cellular immune system for captive mammals, it is yet poorly understood how ecological factors are connected with the different branches of the immune system in wild populations. Here, we tested how variation in aspects of the constitutive cellular and humoral immune system of free ranging bats is associated with two ecological factors that likely influence the putative risk of species to become infected by parasites and pathogens: diet and shelter. We found that white blood cell counts of 24 syntopic Neotropical bat species varied with the species' diet and body mass. Bats that included at least partially vertebrates in their diet exhibited the highest white blood cell counts, followed by phytophagous and insectivorous species, which is in agreement with the assumption that the immune system varies with the pathogen transmission risk of a trophic level. The soluble part of the constitutive immune response, assessed by an in vitro bacterial killing assay, decreased with increasing roost permanence. Our results suggest that the ecology is an important factor in the evolution of the immune system in bats and probably also other mammals. Citation: Schneeberger K, Czirják GÁ , Voigt CC (2013) Measures of the Constitutive Immune System Are Linked to Diet and Roosting Habits of Neotropical Bats. PLoS ONE 8(1): e54023.

Adaptive immunity allows vertebrates to gain protection against repeated pathogenic infections. Analogous responses (priming) have been recently uncovered in invertebrates. However, whether such responses are widespread is not known. The present study investigated the presence of immunological priming in males of a species whose phylogenetic position places it in one of the less derived insect orders. It is hypothesized that the efficiency of such a response could be related to animal condition, as assessed by the expression of a sexually selected ornament. Hetaerina americana Fabricius (Odonata: Calopterygidae) males bear a conspicuous ornament (a red wing spot), which is evolutionarily maintained via male territorial competition. Using field-collected animals, a group of males is challenged with bacteria before exposure to a higher dose of the same or a different bacteria, and survival is compared with that of infected males not previously challenged, as well as control groups. Grampositive and Gram-negative bacteria are used. To explore how long priming may take to work, the second exposure is carried out either after 1 or 5 days. Red spot and body size are entered in the analysis as predictors of survival within and between groups. There is no difference in survival among groups, which suggests no priming effect. Overall, red spot and body size are not consistent in explaining survival.

Adaptive immunity allows vertebrates to gain protection against repeated pathogenic infections. Analogous responses (priming) have been recently uncovered in invertebrates. However, whether such responses are widespread is not known. The present study investigated the presence of immunological priming in males of a species whose phylogenetic position places it in one of the less derived insect orders. It is hypothesized that the efficiency of such a response could be related to animal condition, as assessed by the expression of a sexually selected ornament. Hetaerina americana Fabricius (Odonata: Calopterygidae) males bear a conspicuous ornament (a red wing spot), which is evolutionarily maintained via male territorial competition. Using field-collected animals, a group of males is challenged with bacteria before exposure to a higher dose of the same or a different bacteria, and survival is compared with that of infected males not previously challenged, as well as control groups. Grampositive and Gram-negative bacteria are used. To explore how long priming may take to work, the second exposure is carried out either after 1 or 5 days. Red spot and body size are entered in the analysis as predictors of survival within and between groups. There is no difference in survival among groups, which suggests no priming effect. Overall, red spot and body size are not consistent in explaining survival.

Adaptive immunity allows vertebrates to gain protection against repeated pathogenic infections. Analogous responses (priming) have been recently uncovered in invertebrates. However, whether such responses are widespread is not known. The present study investigated the presence of immunological priming in males of a species whose phylogenetic position places it in one of the less derived insect orders. It is hypothesized that the efficiency of such a response could be related to animal condition, as assessed by the expression of a sexually selected ornament. Hetaerina americana Fabricius (Odonata: Calopterygidae) males bear a conspicuous ornament (a red wing spot), which is evolutionarily maintained via male territorial competition. Using field-collected animals, a group of males is challenged with bacteria before exposure to a higher dose of the same or a different bacteria, and survival is compared with that of infected males not previously challenged, as well as control groups. Grampositive and Gram-negative bacteria are used. To explore how long priming may take to work, the second exposure is carried out either after 1 or 5 days. Red spot and body size are entered in the analysis as predictors of survival within and between groups. There is no difference in survival among groups, which suggests no priming effect. Overall, red spot and body size are not consistent in explaining survival.

Parasites are characteristically aggregated within hosts, but identifying the mechanisms underlying such aggregation can be difficult in wildlife populations. We examined the influence of host age and sex over an annual cycle on the eggs per gram of feces (EPG) of nematode parasites infecting wild Japanese macaques (Macaca fuscata yakui) on Yakushima Island. Five species of nematode were recorded from 434 fecal samples collected from an age-structured group of 50 individually recognizable macaques. All parasites exhibited aggregated EPG distributions. The age-infection profiles of all three directly transmitted species (Oesophagostomum aculeatum, Strongyloides fuelleborni, and Trichuris trichiura) exhibited convex curves, but concavity better characterized the age-infection curves of the two trophically transmitted species (Streptopharagus pigmentatus and Gongylonema pulchrum). There was a male bias in EPG and prevalence of infection with directly transmitted species, except in the prevalence of O. aculeatum, and no sex bias in the other parasites. Infection with O. aculeatum showed a female bias in prevalence among young adults, and additional interactions with sex and seasonality show higher EPG values in males during the mating season (fall) but in females during the birth season (spring). These patterns suggest that an immunosuppressive role by reproductive hormones may be regulating direct, but not indirect, lifecycle parasites. Exposure at an early age may trigger an immune response that affects all nematodes, but trophically transmitted species appear to accumulate thereafter. Although it is difficult to discern clear mechanistic explanations for parasite distributions in wildlife populations, it is critical to begin examining these patterns in host species that are increasingly endangered by anthropogenic threats.