Individual differences in peripheral blood immunological and hormonal measures in adult male rhesus macaques (Macaca mulatta): Evidence for temporal and situational consistency

A growing body of research has indicated that consistent individual differences exist in physiological systems with which the immune system interacts. Few data have been reported that demonstrate stable individual differences in immunological measures, however. In the present study, enumerative measures of immune system activity were examined in 36 adult male rhesus macaques over a 13 month period under baseline conditions as well as under conditions of pharmacological and physical challenge. Blood samples were assayed for plasma concentrations of ACTH and cortisol, as well as neutrophil, total lymphocyte, CD4+ and CD8+ lymphocyte numbers, and the CD4/CD8 ratio. Analyses revealed that individual differences in the CD4/CD8 ratio and, to a lesser extent, plasma ACTH and cortisol concentrations, and neutrophil and CD4+ lymphocyte numbers were consistent across situations and times, despite changes in mean values during the various blood sampling sessions. The results suggest that the CD4/CD8 ratio might be considered trait-like and a useful immunological measure of biobehavioral organization. Am. J. Primatol. 44:29–41, 1998. © 1998 Wiley-Liss, Inc.     

Interaction between sexual steroids and immune response in affecting oxidative status of birds

One hypothesis explaining the honesty of secondary sexual traits regulated by testosterone (T) is that T can impair the balance between pro-oxidant compounds and antioxidant defences, favouring a status of oxidative stress that only good quality individuals can sustain (oxidative handicap hypothesis). In the present study, we evaluated for the first time the effects of sexual steroids, T and its metabolites 5-α-dihydrotestosterone (DHT) and estradiol (E2) on oxidative damage and plasma non-enzymatic antioxidant capacity, while birds are faced by an oxidative challenge induced by an immune stimulation with sheep red blood cells. We used male and female diamond doves Geopelia cuneata, a species that shows an orange-red periorbital ring, whose size and color are strongly affected by androgens, but not by estrogens. Immunization increased oxidative damage in all groups, regardless of hormone treatment. It also decreased anti-oxidant capacity in all groups, except for testosterone treated birds. The ratio of oxidative damage over anti-oxidant capacity (oxidative stress) was increased in both immunological challenged controls and E2 birds, while challenged birds treated with androgens did not differ from non-challenged birds. The response of males and females to our treatments never differed. Our results undermine the idea that T can induce honest signalling through a pro-oxidant activity.     

Growing old with the immune system: a study of immunosenescence in the zebra finch (<Emphasis Type="Italic">Taeniopygia guttata)</Emphasis>

Immunosenescence has not received much attention in birds and the few existing studies indicate that the occurrence of immunosenescence and/or its extent may differ between species. In addition, not much information is available on the immunosenescence patterns of different immune parameters assessed simultaneously in both sexes within a single species. The present study reports the results on immunosenescence in innate immunity and both cellular and humoral acquired immunity of both sexes in a captive population of zebra finch (Taeniopygia guttata) using three age groups (approximately 0.2, 2.5 and 5.1 years). Both male and female finches showed an inverse U-shaped pattern in cellular immune function with age, quantified by a PHA response. Males showed stronger responses than females at all ages. In contrast, an increase with age in humoral immunity, quantified through total plasma immunoglobulin Y levels, was found in both sexes. However, our measurements of innate immunity measured through the bacteria-killing ability against Escherichia coli gave inconclusive results. Still, we conclude that both cellular and humoral acquired immunity are susceptible to immunosenescence, and that the sexes differ in cellular immunity.     

Measuring circulating antioxidants in wild birds

Antioxidants protect against free radical damage, which is associated with various age-related pathologies. Antioxidants are also an important buffer against the respiratory burst of the immune system. This protection presumably has costs and therefore might underlie important life-history trade-offs. Studying such trade-offs in a comparative context requires field-applicable methods for assessing antioxidant capacity in wild animals. Here, we present modifications to a simple spectrophotometric assay (the TEAC or TAS assay) that can be applied to miniscule amounts of blood plasma to determine circulating antioxidant capacity. Additionally, uric acid, the most abundant circulating antioxidant, should be measured independently. Uric acid in birds is derived from amino acid catabolism, perhaps incidentally to its antioxidant function. The assay was validated in experimental studies on chickens showing effects of diet on antioxidant capacity, and in field measurements on 92 species of birds, which demonstrate substantial species differences in constitutive antioxidant capacity. Furthermore, most wild birds demonstrate a dramatic change in antioxidant capacity due to stress. These results show that this technique detects variation appropriate for both interspecific and intraspecific studies, and that antioxidants and uric acid change in response to conditions of interest to field ecologists, such as diet and stress.     

Assessing immunological function in toxicological studies of avian wildlife

Laboratory and field studies have demonstrated that the immunesystem is sensitive to environmental contaminants. Testing protocolshave been developed to screen for immunotoxic effects and elucidatemechanisms of toxicity in laboratory rodents. Similar methodshave been applied to wildlife species in captivity and the wild.Several epizootics in wildlife have been associated with elevatedexposure to contaminants. This paper discusses immunotoxicologicaltechniques used in studies of avian wildlife. Measurements ofimmunological structure include peripheral white blood cellcounts and the mass and cellularity of immune organs such asthe thymus, spleen, and bursa of Fabricius. While contaminantscan alter these measures of immunological structure, such measuresdo not directly assess how the immune system functions, i.e.,responds to specific challenges. The two most commonly usedin vivo immune function tests in birds are the phytohemagglutinin(PHA) skin response for T cell-mediated immunity and the sheepred blood cell (SRBC) hemagglutination assay for antibody-mediatedimmunity. In vitro tests of immune function in avian wildlifeinclude proliferation of lymphocytes in response to variousmitogens and phagocytosis of fluorescent particles by monocytes.While optimization of in vitro techniques for wildlife speciesis often time-consuming, these assays usually require only asingle blood sample and can elucidate mechanisms of toxicity.In immunological studies of wildlife, investigators should considerfactors that may influence immune responses, including age,body condition, date, developmental stage of the immune system,and time required for the progression of immune responses.     

Effects of corticosterone treatment on growth, development, and the corticosterone response to handling in young Japanese quail (Coturnix coturnix japonica)

Corticosterone, a glucocorticoid secreted during stress responses, has a range of actions that help birds respond to stressors. Although effects of corticosterone treatment have been described in several avian species, the impacts of defined increases in plasma corticosterone on early development and on corticosterone stress responses are little known. These issues were addressed by providing quail with different doses of corticosterone in drinking water from days 8 to 38 post-hatch. The corticosterone dose consumed by each bird during treatment days 15-30 was calculated by measuring water intake. The corticosterone dose was inversely, but weakly, correlated with weights of the bursa, thymus, spleen, liver, testes, oviduct, muscle, and body, and positively correlated with peritoneal fat deposition. When birds were divided into groups based on their corticosterone intake, weights of the spleen, thymus, bursa, muscle, testes, and oviduct were significantly reduced in birds receiving the highest doses; with the exception of muscle, similar reductions were also observed in birds receiving medium doses, and thymic growth was inhibited in birds receiving low doses. The acute corticosterone stress response was measured by handling birds for 15 min. Plasma corticosterone was transiently increased at 15 min in control birds in response to the handling stressor. Some birds consuming low doses of corticosterone had corticosterone responses similar to control birds. Initial corticosterone concentrations were elevated in birds consuming higher doses of corticosterone. Plasma corticosterone in these birds decreased from 0 to 15 min, then increased from 15 to 30 min. The initial decrease could be due to corticosterone clearance, whilst the increase could indicate that the birds had a greater response than control birds to isolation as a stressor. Corticosterone treatment may have reduced the strength of corticosterone negative feedback within the hypothalamo-pituitary-adrenal axis. The results indicate that individuals and organs differ in their sensitivity to corticosterone. Moreover, elevated plasma corticosterone may disrupt the acute corticosterone stress response, and may thus reduce the ability of birds to cope with stressors.     

Plasma reactive oxygen metabolites and non-enzymatic antioxidant capacity are not affected by an acute increase of metabolic rate in zebra finches

Understanding the sources of variation in oxidative stress level is a challenging issue due to the implications of oxidative stress for late age diseases, longevity and life-history trade-offs. Reactive oxygen species that cause oxidative stress are mostly a by-product of energy metabolism and it is therefore often assumed that oxidative stress is proportional to energy consumption. In mammals, an increased metabolic rate induced by cold exposure generally increases oxidative stress. However, compared to mammals, birds generate fewer free radicals per ATP produced and hence it is not obvious that, in birds, a cold-induced increase of metabolic rate increase oxidative stress. We tested whether cold-induced increase in metabolic rate increased oxidative stress in zebra finches by exposing individuals to cold and warm overnight temperatures. We registered metabolic rate and plasma levels of non-enzymatic antioxidants and reactive oxygen metabolites (ROMs), a measure of oxidative damage. Metabolic rate was on average 88 % higher in cold compared to warm temperature, with females being stronger affected than males. However, temperature had no effect on plasma antioxidants or our measure of oxidative damage. Middle-age birds had higher levels of plasma antioxidants than younger and older birds, but age was unrelated to ROMs. Birds showed repeatability of plasma ROMs across temperatures but not of non-enzymatic antioxidants. In contrast to similar studies in mammals, our results do not show evidence of increased oxidative stress in plasma after an acute cold-induced increase of metabolic rate but research in more bird species is needed to assess the generality of this pattern.     

Immune indexes of larks from desert and temperate regions show weak associations with life history but stronger links to environmental variation in microbial abundance

Abstract Immune defense may vary as a result of trade-offs with other life-history traits or in parallel with variation in antigen levels in the environment. We studied lark species (Alaudidae) in the Arabian Desert and temperate Netherlands to test opposing predictions from these two hypotheses. Based on their slower pace of life, the trade-off hypothesis predicts relatively stronger immune defenses in desert larks compared with temperate larks. However, as predicted by the antigen exposure hypothesis, reduced microbial abundances in deserts should result in desert-living larks having relatively weaker immune defenses. We quantified host-independent and host-dependent microbial abundances of culturable microbes in ambient air and from the surfaces of birds. We measured components of immunity by quantifying concentrations of the acute-phase protein haptoglobin, natural antibody-mediated agglutination titers, complement-mediated lysis titers, and the microbicidal ability of whole blood. Desert-living larks were exposed to significantly lower concentrations of airborne microbes than temperate larks, and densities of some bird-associated microbes were also lower in desert species. Haptoglobin concentrations and lysis titers were also significantly lower in desert-living larks, but other immune indexes did not differ. Thus, contrary to the trade-off hypothesis, we found little evidence that a slow pace of life predicted increased immunological investment. In contrast, and in support of the antigen exposure hypothesis, associations between microbial exposure and some immune indexes were apparent. Measures of antigen exposure, including assessment of host-independent and host-dependent microbial assemblages, can provide novel insights into the mechanisms underlying immunological variation.     

Effects of housing conditions during the rearing and laying period on adrenal reactivity, immune response and heterophil to lymphocyte (H/L) ratios in laying hens

This study was conducted to evaluate the effect of early rearing conditions on physiological, haematological and immunological responses relevant to adaptation and long-term stress in white Leghorn hens with intact beaks housed in furnished cages (FC) or conventional cages (CC) during the laying period. Pullets were cage reared (CR) or litter floor reared (FR). From 16 to 76 weeks of age, hens were housed in FC (eight hens per cage) or in CC (three hens per cage). As measures of long-term stress at the end of the laying period, adrenal reactivity was quantified by assessing corticosterone responses to adrenocorticotropin challenge, and immune response was assessed by measuring antibody responses after immunization with sheep red blood cells (SRBC) and keyhole limpet haemocyanin (KLH). Heterophil to lymphocyte (H/L) ratio was employed as an indicator of stress. Rearing conditions significantly affected anti-SRBC titres (P < 0.0001) and tended to affect H/L ratios (P = 0.07), with the highest values found in FR hens. Layer housing affected H/L ratio (P < 0.01); the highest ratio was found in FR birds housed in FC during the laying period. This study shows that early rearing environment affects immunological indicators that are widely used to assess stress in laying hens. However, while results on H/L ratio indicated that FR birds experienced more stress particularly when they were housed in FC during the laying period, the immune responses to SRBC in FR hens was improved, indicating the opposite. This contradiction suggests that the effects on immune response may have been associated with pathogenic load due to environmental complexity in FR and FC hens rather than stress due to rearing system or housing system per se.     

Interspecific variation in avian blood parasites and haematology associated with urbanization in a desert habitat

Many avian species are negatively impacted by urbanization, but other species survive and prosper in urbanized areas. One factor potentially contributing to the success of some species in urban areas is the reduced presence of predators or parasite vectors in urban compared to rural areas. In addition, urban areas may provide increased food and water resources, which can enhance immune capacity to resist infection and the ability to eliminate parasites. We determined patterns of blood parasitism, body condition, and immune cell profiles in urban and rural populations of five adult male songbird species that vary in their relative abundance within urban areas. Urban birds generally exhibited less blood parasitism than rural birds. This difference was particularly evident for the urban-adaptable Abert's towhee Pipilo aberti . In contrast, no difference in haemoparasitism was seen between urban and rural populations of the curve-billed thrasher Toxostoma curvirostre , a less-urban adaptable species. In two closely related species, the curve-billed thrasher and the northern mockingbird Mimus polyglottos , urban birds had a higher leukocyte count and a higher heterophil to lymphocyte ratio, which is often associated with chronic stress or current infection, than rural birds. Urban northern mockingbirds were in better condition than rural counterparts, but no habitat-related differences in condition were detected for other species. Parasitic infection was correlated with body condition in only one species, the canyon towhee Pipilo fuscus . Parasitic infection in most species was correlated with changes in leukocyte abundance and profile. The findings suggest that interspecific differences in parasitic infection cannot be attributed entirely to differences in vector abundance or body condition. Interactions between immune function, parasite infection risk, and resource availability may contribute to determining the relative ability of certain species to adapt to cities.     

Sarcocystis transmitted by blood transfusion.

Merozoites were found in blood smears from calves, lambs, and pigs exhibiting signs of acute sarcocystosis after oral infection with sporocysts of Sarcocystis bovicanis, Sarcocystis ovicanis, and Sarcocystis suihominis. In each of 3 experiments, whole blood containing merozoites was transfused from an infected host to 2 uninfected recipients of the same species; 2 additional animals of the same species served as uninfected nontransfused controls. The bovine, ovine, and porcine donors all died of acute sarcocystosis. Clinical signs of sarcocystosis were not seen in any transfusion recipients or controls. All recipients and controls were killed and histologic specimens of esophagus, diaphragm, heart, skeletal muscle, and tongue were examined microscopically. Large numbers of intramuscular cysts were present in transfusion recipients, whereas few or no cysts were present in controls, indicating that S. bovicanis, S. ovicanis, and S. suihominis had been transmitted between intermediate hosts of the same species by blood transfusion.     

Constitutive innate immunity is a component of the pace-of-life syndrome in tropical birds

We studied the relationship between one component of immune function and basal metabolic rate (BMR), an indicator of the 'pace-of-life syndrome', among 12 tropical bird species and among individuals of the tropical house wren (Troglodytes aedon), to gain insights into functional connections between life history and physiology. To assess constitutive innate immunity we introduced a new technique in the field of ecological and evolutionary immunology that quantifies the bactericidal activity of whole blood. This in vitro assay utilises a single blood sample to provide a functional, integrated measure of constitutive innate immunity. We found that the bactericidal activity of whole blood varied considerably among species and among individuals within a species. This variation was not correlated with body mass or whole-organism BMR. However, among species, bacteria killing activity was negatively correlated with mass-adjusted BMR, suggesting that species with a slower pace-of-life have evolved a more robust constitutive innate immune capability. Among individuals of a single species, the house wren, bacteria killing activity was positively correlated with mass-adjusted BMR, pointing to physiological differences in individual quality on which natural selection potentially could act.     

Normal levels of immunocompetence in possums (Trichosurus vulpecula) exposed to different laboratory housing conditions post capture

Specific and non-specific immunological tests were used to monitor aspects of the immune response in captive possums. The tests included total and differential white blood cell counts, lymphocyte transformation assay, and enzyme linked immunosorbant assay. The level of free cortisol present in possum plasma samples was evaluated as an endocrine marker for stress. Four different housing conditions were used to test whether stress could be managed or avoided in captive animals. Animals were caged individually or as groups in pens. Bacille Calmette-Gurein (BCG) and tetanus toxoid immunization was used to evoke primary cell mediated and antibody responses in test animals. The results indicated that there was no significant difference in immunological responses or endocrine parameters in animals held under any of the housing conditions. The results infer that wild possums adapt quickly post-capture to novel housing conditions and produce representative patterns of immunity when held in housing conditions and fed ad libitum.     

Immune responses in Rhodnius prolixus: influence of nutrition and ecdysone

Starved larvae of Rhodnius prolixus, when challenged with Enterobacter cloacae B12, had their mortality related to their period of starvation. R. prolixus larvae fed on plasma alone, compared with insects fed on whole blood, had their immune reactivity affected as shown by: (i) a significant reduction in the ability to produce cecropin-like and lysozyme activities in the haemolymph when inoculated with E. cloacae; (ii) a reduction in numbers of haemocytes and nodule formation following challenge with bacteria; (iii) a decreased ability of plasma-fed insects in destroying their infection caused by inoculation of E. cloacae cells; and (iv) alpha-ecdysone therapy counteracted the immune depression in Rhodnius larvae fed on plasma alone. However, unlike other immune reactions, this set of experiments failed to demonstrate any interference of the plasma feeding on the prophenoloxidase-activating system, since melanin production was not reduced when the system was stimulated by the presence of bacteria in the haemolymph. The significance of these data is discussed in relation to the effect of diet components and the moulting hormone on the immune reactivity in insects.     

No simple answers for ecological immunology: relationships among immune indices at the individual level break down at the species level in waterfowl

Understanding immune function in the context of other life-history traits is crucial to understand the evolution of life histories, at both the individual and species levels. As the interest in assessing immune function for these comparative purposes grows, an important question remains unanswered: can immune function be broadly characterized using one or two simple measures? Often, interpretation of individual assays is ambiguous and relationships among different measures of immune function remain poorly understood. Thus, we employed five protocols to measure 13 variables of immune function in ten species of waterfowl (Anseriformes). All assays were based on a single blood sample subdivided into leukocyte (blood smear) and plasma (frozen until analysis) components. All assays were run using samples from every individual, and a nested analysis was used to partition variation/covariation at the levels of species and individuals within species. We detected positive correlations between functionally related measures of immunity within species, but these were absent from comparisons between species. A canonical correlation analysis revealed no significant relationships between the plasma and leukocyte assays at the levels of both individual and species, suggesting that these measures of immunity are neither competitive nor synergistic. We conclude that one measure of each assay type may be required to maximally characterize immune function in studies of a single species, while the same is not true in studies among species.     

Diving behaviour of white-chinned petrels and its relevance for mitigating longline bycatch

The white-chinned petrel (Procellaria aequinoctialis) is the seabird species most commonly killed by Southern Hemisphere longline fisheries. Despite the importance of diving ability for mitigating longline bycatch, little is known of this species’ diving behaviour. We obtained data from temperature–depth recorders from nine white-chinned petrels breeding on Marion Island, southwestern Indian Ocean, during the late incubation and chick-rearing period. Maximum dive depth (16 m) was slightly deeper than the previous estimate (13 m), but varied considerably among individuals (range 2–16 m). Males dived deeper than females, and birds feeding chicks dived deeper than incubating birds, but dive rate did not differ between the sexes. Time of day had no significant effect on dive depth or rate. Our findings will help to improve the design and performance of mitigation measures aimed at reducing seabird bycatch in longline fisheries, such as the calculation of minimum line sink rates and optimum aerial coverage of bird-scaring lines.     

Sex-specific variation in brown-headed cowbird immunity following acute stress: a mechanistic approach

There is some discrepancy in the literature regarding whether acute stress is immunostimulatory or immunosuppressive. Studies of domesticated (laboratory and food) animals and humans typically indicate that acute stress is immunostimulatory, whereas studies of non-domesticated species document both immunostimulatory and immunosuppressive results. Few studies have examined the mechanisms responsible for changes in immune activity in species other than those classically used in laboratory research. We examined the effect of both acute stress and exogenous corticosterone (CORT) on the bactericidal capacity (BC) of blood plasma from captive, wild-caught brown-headed cowbirds (Molothrus ater) to determine if CORT is responsible for changes in levels of immune activity. We conducted "stress tests" in which we handled birds to elicit a stress response and then measured the birds' total CORT and BC at 30 or 90 min post-stressor. We also conducted non-invasive tests in which we administered exogenous CORT by injecting it into mealworms that were fed to the cowbirds remotely. Total, free, and bound CORT levels, corticosteroid binding globulins (CBGs), and BC at 7 or 90 min post-mealworm ingestion were measured. Both males and females exhibited significant increases in total CORT following handling stress and the administration of exogenous CORT. Experimental males and females also exhibited a significant increase in CBG capacity at 7 min post-mealworm ingestion compared to controls. Male cowbirds exhibited a significant decline in their BC following both handling stress and the administration of exogenous CORT whereas female cowbirds exhibited no decline under either condition. Female CBG levels were not different than those of males, suggesting that differences in BC could be due to differences between the sexes in the number of corticosteroid receptors which, along with CBGs, regulate the stress response. Female cowbirds may modulate their stress response as an adaptive life-history strategy for maximizing current reproduction.     

Coccidian infection causes oxidative damage in greenfinches

The main tenet of immunoecology is that individual variation in immune responsiveness is caused by the costs of immune responses to the hosts. Oxidative damage resulting from the excessive production of reactive oxygen species during immune response is hypothesized to form one of such costs. We tested this hypothesis in experimental coccidian infection model in greenfinches Carduelis chloris. Administration of isosporan coccidians to experimental birds did not affect indices of antioxidant protection (TAC and OXY), plasma triglyceride and carotenoid levels or body mass, indicating that pathological consequences of infection were generally mild. Infected birds had on average 8% higher levels of plasma malondialdehyde (MDA, a toxic end-product of lipid peroxidation) than un-infected birds. The birds that had highest MDA levels subsequent to experimental infection experienced the highest decrease in infection intensity. This observation is consistent with the idea that oxidative stress is a causative agent in the control of coccidiosis and supports the concept of oxidative costs of immune responses and parasite resistance. The finding that oxidative damage accompanies even the mild infection with a common parasite highlights the relevance of oxidative stress biology for the immunoecological research.     

The effect of heat stress on intestinal integrity and Salmonella invasion in broiler birds

The intestinal mucosa works as a barrier to protect the internal environment of the animal from bacteria and bacterial toxins found in the gut lumen. Heat stress may harm this function. Therefore, we designed the current experiment to investigate the effect of heat stress on intestinal integrity, physiological and immunological responses and Salmonella invasion in broiler chickens. At 26 days of age, 72 birds were randomly distributed into 3 treatments, with 8 replicates per treatment and 3 birds per replicate. The three treatments were control treatment; kept at thermoneutral environmental conditions (20 ± 2 °C), chronic heat stress treatment (exposed to 30 ± 2 °C; 24 h/day) and acute heat stress treatment (exposed to 35 ±2 °C from 09:00 to 13:00 and kept at 20 ± 1 °C from 13:00 to 09:00). The heat stress exposure was conducted for 10 successive days. Compared with the control treatment, birds subject to chronic and acute heat stress had reduced (P < 0.05) body weight and body gain and increased (P < 0.05) feed conversion ratio. However, feed intake and mortality rate were only increased (P < 0.05) in the acute heat stress treatment. Rectal temperature and Δ rectal temperature (°C/h) increased (P < 0.05) sharply during the first 2 days of exposure followed by gradual decreases until a plateau was achieved. Heat-stressed birds had increased (P < 0.05) serum concentrations of corticosterone, endotoxin lipopolysaccharide and the systemic inflammatory cytokine: TNF-α and IL-2, as well as a higher (P < 0.05) prevalence of Salmonella spp. in meat and livers, as compared with control treatment. It can be concluded that heat stress impaired intestinal integrity which resulted in increased intestinal permeability to endotoxin, translocation of intestinal pathogens (Salmonella spp.) and serum inflammatory cytokines. Therefore, avoiding thermal dysfunction of intestinal barrier is a significant factor in maintaining welfare, immune status and meat safety of broiler birds.