Secondary phytohaemagglutinin (PHA) swelling response is a good indicator of T‐cell‐mediated immunity in free‐living birds

The validity of using the phytohaemagglutinin (PHA) test to measure acquired immunity, one of the most widely used methods, is currently being debated due to new knowledge on the complex physiology of the process. As a greater secondary response to repeated challenges linked to increases of circulating lymphocyte levels would be indicative of a T‐cell‐mediated immune response, we performed for the first time an experiment under natural conditions with repeated PHA challenges in free‐living adult birds and chicks to shed light on this topic. We found significantly stronger secondary response to PHA injection independent of sex or age, while controlling for body condition, the second response being on average 90% larger than the first. Likewise, lymphocyte counts were significantly higher in the second PHA challenge, whereas no significant differences were found among untreated birds. Significant positive correlations between the PHA response and both lymphocyte counts and plasma protein levels (mainly albumin, globulin precursor) were recovered, whereas no significant differences were recovered in plasma protein levels between challenges. Our results are consistent with those from captive birds, supporting the validity of the PHA skin‐swelling test as an accurate gauge of acquired T‐cell‐mediated immunity in birds.     

Experimentally elevated testosterone levels enhance courtship behaviour and territoriality but depress acquired immune response in Red Bishops Euplectes orix

The immunocompetence handicap hypothesis (ICHH) posits that the hormone testosterone mediates a trade‐off between investment in reproduction and immunological condition. In this study, we tested the ICHH in the Red Bishop Euplectes orix, a polygynous weaverbird. Males of this strongly sexually dimorphic species show an elaborate courtship display to attract females and compete aggressively with other males for nesting sites in breeding colonies. We experimentally elevated testosterone levels in breeding male Red Bishops kept in an aviary with a subcutaneous implantation of testosterone‐releasing pellets. We then compared behaviour, development of territory size and immunological condition (as assessed through a white blood cell count) of the experimental group with a control group treated with placebos. In addition, we measured the primary and secondary response to phytohaemagglutinin (PHA) to investigate the effect of testosterone on both innate and acquired immunity. Males with elevated levels of testosterone enlarged their territories and conducted more courtship behaviour, while showing a decrease in health, expressed by an increased heterophil/lymphocyte ratio compared with the control group. Males of the control group showed an increase of the secondary response to PHA, as expected under the assumption that repeated exposure to an antigen enhanced the immune response due to acquired immunity. However, males with experimentally increased testosterone levels did not show such an enhanced immune response in the secondary PHA test (although sample size and power of the statistical tests were low), indicating that testosterone treatment might directly or indirectly suppress some component of the acquired immune response.     

The PHA Test as an Indicator of Phagocytic Activity in a Passerine Bird

Several techniques in ecological immunology have been used to assess bird immunocompetence thus providing useful information to understand the contribution of the immunological system in life-history decisions. The phytohaemagglutinin (PHA)-skin test has been the most widely employed technique being interpreted as the sole result of T lymphocytes proliferation and hence used to evaluate acquired immunological capacity. However, the presence of high numbers of phagocytic cells in the swelling point has cast some doubt about such an assumption. To address this issue, we collected blood from 14 days-old nestlings of spotless starling (Sturnus unicolor), administered subcutaneous PHA immediately after and then measured the swelling response 24 hours later. Differential counts of white blood cells suggested that an intense development of acquired immunological defences was taking place. The phagocytic activity of both heterophiles and monocytes was also very intense as it was the swelling response. Moreover, our results show, for the first time in birds, a positive relationship between the phagocytic activity of both kinds of cells and the swelling response. This broadens the significance of the PHA test from reflecting T lymphocytes proliferation -as previously proposed but still undetermined in vivo- to evaluate phagocytosis as well. In other words, our data suggest that the PHA swelling response may not be considered as the only consequence of processes of specific and induced immunity –T lymphocytes proliferation- but also of constitutive and nonspecific immunity –heterophiles and monocytes phagocytosis. We propose the extensive use of PHA-skin test as an optimal technique to assess immunocompetence.     

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.     

Phytohemagglutinin response and immunoglobulin index decrease during incubation fasting in female common eiders

To maximize their fitness, long-lived species face trade-offs between survival and reproduction. The cost of reproduction, which is defined as the negative effect of current parental investment on chances of adult survival and future reproduction, may affect immune function, possibly through hormonal changes. In this study, components of acquired immunity and plasma corticosterone levels of female eiders (Somateria mollissima) have been measured throughout the incubation period as a function of clutch size. These precocial birds lay up to six eggs and fast completely during incubation. Birds were sampled early and late in the incubation period, with clutches ranging from one to four eggs. T-cell-mediated immune response and humoral immunity were assessed by phytohemagglutinin (PHA) skin tests (a challenging method) and measurements of serum immunoglobulins (a monitoring method), respectively. During incubation, responses to PHA injection and immunoglobulin index significantly decreased, by about 40% and 25%, respectively. These observed decreases occurred independently of the number of eggs laid by the females. Corticosterone did not vary significantly during incubation, whatever the clutch size. We conclude that female eiders seem to reallocate their resources from immune function to reproductive effort independently of clutch size or corticosterone levels.     

Mosquito immune responses and malaria transmission: lessons from insect model systems and implications for vertebrate innate immunity and vaccine development

The introduction of novel biochemical, genetic, molecular and cell biology tools to the study of insect immunity has generated an information explosion in recent years. Due to the biodiversity of insects, complementary model systems have been developed. The conceptual framework built based on these systems is used to discuss our current understanding of mosquito immune responses and their implications for malaria transmission. The areas of insect and vertebrate innate immunity are merging as new information confirms the remarkable extent of the evolutionary conservation, at a molecular level, in the signaling pathways mediating these responses in such distant species. Our current understanding of the molecular language that allows the vertebrate innate immune system to identify parasites, such as malaria, and direct the acquired immune system to mount a protective immune response is very limited. Insect vectors of parasitic diseases, such as mosquitoes, could represent excellent models to understand the molecular responses of epithelial cells to parasite invasion. This information could broaden our understanding of vertebrate responses to parasitic infection and could have extensive implications for anti-malarial vaccine development.     

Plant innate immunity: An updated insight into defense mechanism

Plants are invaded by an array of pathogens of which only a few succeed in causing disease. The attack by others is countered by a sophisticated immune system possessed by the plants. The plant immune system is broadly divided into two, viz. microbial-associated molecular-patterns-triggered immunity (MTI) and effector-triggered immunity (ETI). MTI confers basal resistance, while ETI confers durable resistance, often resulting in hypersensitive response. Plants also possess systemic acquired resistance (SAR), which provides long-term defense against a broad-spectrum of pathogens. Salicylic-acid-mediated systemic acquired immunity provokes the defense response throughout the plant system during pathogen infection at a particular site. Trans-generational immune priming allows the plant to heritably shield their progeny towards pathogens previously encountered. Plants circumvent the viral infection through RNA interference phenomena by utilizing small RNAs. This review summarizes the molecular mechanisms of plant immune system, and the latest breakthroughs reported in plant defense. We discuss the plant–pathogen interactions and integrated defense responses in the context of presenting an integral understanding in plant molecular immunity.     

Ecological and evolutionary implications of immunological priming in invertebrates

Invertebrates have an immune response that differs considerably from the acquired immune response found in vertebrates. However, new studies indicate that past experience with a pathogen can provide individual invertebrates, or their descendants, with enhanced immunity. This prophylactic effect, termed immunological priming, is functionally similar to the acquired immune response in vertebrates. This newfound complexity of invertebrate immunity begs investigation into the conditions under which immunological priming should evolve, and its consequences for population dynamics.     

禁食时间对雌性长爪沙鼠免疫功能的影响

动物的免疫系统是防御细菌、病毒等病原体侵害的重要途径,对其生存很重要,而食物短缺是影响动物免疫的重要因素。前期研究发现,禁食3天可抑制雌性长爪沙鼠T细胞介导的免疫功能,而禁食时长可能在影响动物免疫中起重要作用。为此,本实验主要测定不同的禁食时间如何影响雌性长爪沙鼠的免疫功能,该问题的解决有助于理解它们在野外面临食物短缺时的生存状况。将55只成年雌鼠随机分为8组,分别禁食1 d、2 d 、3 d和自由取食,每种处理又分别注射磷酸缓冲液和植物血凝素（PHA）。结果发现,随禁食时间的延长,PHA反应（指示T细胞介导的免疫功能）逐渐受到抑制,胸腺和脾脏逐渐萎缩,表明禁食时间对长爪沙鼠的免疫功能影响显著。同样,随饥饿时间增加,长爪沙鼠的体脂、血糖和血清瘦素水平逐渐下降,并且与PHA反应呈正相关,但白细胞总数和皮质酮水平不受禁食时间的影响。结果表明,禁食时间长短能显著影响雌性长爪沙鼠的免疫功能,随禁食时间的延长,长爪沙鼠T细胞介导的免疫功能逐渐受抑制,这可能与体脂和葡萄糖含量下降有关,血清瘦素逐渐降低也可能是其免疫受抑制的原因。     

Mycobacterial heat shock proteins as vaccines - a model of facilitated antigen presentation

Heat shock proteins (hsps) are a highly conserved family of proteins, first recognized by their upregulated expression in response to host exposure to raised temperatures. Further study has revealed that they have numerous functions in the cell, primarily as chaperones mediating both the correct folding of nascent polypeptide chains and the dissolution of aggregated protein complexes. The energy requirement for this chaperone activity is provided by the ATPase activity found in most families of hsps and thus the peptide binding capacity is controlled by ATP hydrolysis. The structural consequence of this is that hsps isolated in situ are found complexed to chaperoned peptides (hspCs). Much previous work has implicated hsps in the immune response to pathogens and recent studies have shown that the interaction of hsps with antigen presenting cells, such as dendritic cells (DCs), mediates the integration of the innate and acquired immune responses. This central role for hspCs in immunity is facilitated by their dual function in both innate immunity, with the induction of cytokines and the maturation of DCs mediated by the hsp component, and acquired immunity, with the trafficking of antigens chaperoned in hspCs for antigen presentation by the mature DCs.     

Melanin-based coloration and immunity in polymorphic population of pied flycatcher, <Emphasis Type="Italic">Ficedula hypoleuca</Emphasis>

A specific interest in the persistence of color polymorphism in some populations of birds and other vertebrates is often linked to ideas about the signaling honesty of bright coloration. The evolution of conspicuous ornamentation could be associated with physiological costs including limitations of the immune system. The study of this process is crucial for an understanding of the maintenance of polymorphic coloration. Here we summarized the results of a study of a pied flycatcher population from the Moscow region (Russia) in 2010–2013. We experimentally induced antibody production by injecting sheep red blood cells (SRBC) and inflammatory swelling by injecting phytohaemagglutinin (PHA) after which we estimated the immune response in breeding males. We used leucocytes-to-erythrocytes and heterophils-to-lymphocytes (H/L) ratios as indicators of infectious, inflammatory processes and stress. The results showed that the feeding rates of males treated with SRBC decreased and negatively related to the intensity of their immune responses. Non-molting males of different color types did not significantly differ in antibody production. Among molting breeders, the immune response to SRBC was significantly higher in pale males than in bright ones with rich melanin-based coloration. In contrast to non-molting males, molting pale males had an increased antibody titer after immunization. The lower humoral immune response was associated with the higher H/L stress index before immunization. The change in H/L after immunization positively correlated with the intensity of the humoral immune response. As opposed to humoral immunity, we did not find any significant predictors, including coloration, molt, or their two-way interaction, to explain the variation in cutaneous inflammatory response to PHA. The results suggest that the apparent advantage of a cryptic male phenotype over a conspicuous phenotype occurring in one of two types of immune response has an impact on the maintenance of color polymorphism in the pied flycatcher.     

Autophagy and innate immunity: Triggering,targeting and tuning

Autophagy is a conserved catabolic stress response pathway that is increasingly recognized as an important component of both innate and acquired immunity to pathogens. The activation of autophagy during infection not only provides cell-autonomous protection through lysosomal degradation of invading pathogens (xenophagy), but also regulates signaling by other innate immune pathways. This review will focus on recent advances in our understanding of three major areas of the interrelationship between autophagy and innate immunity, including how autophagy is triggered during infection, how invading pathogens are targeted to autophagosomes, and how the autophagy pathway participates in “tuning” the innate immune response.     

Short-term fluctuations in cellular immunity of tree swallows feeding nestlings

We examined cellular immunity of adult tree swallows feeding nestlings under variable weather conditions. Birds received an injection of phytohaemagglutinin (PHA), which causes a local swelling, reflecting the strength of T-cell-mediated immunocompetence. There was a negative relationship between the immune response and the number of nestlings in the brood (range 3-6 young) which suggests that parental effort suppresses the immune function. However, there was also a strong effect of ambient temperature and food abundance (aerial insects) on immune response. Parents that received the PHA injection during cold weather and at low food abundance showed a suppressed immune response compared to birds treated during more favourable conditions. They also lost more body mass during the 24 h inoculation period, and their offspring showed reduced growth. When controlling for ambient temperature and food abundance in a multivariate analysis, there was no longer any significant effect of brood size on the parents' immune response. Three of 39 pairs deserted their broods after PHA injection. All three desertions took place when the mean ambient temperature fell below 13°C. The PHA response is known to have both heritable and environmental components; our study emphasizes its condition-dependency. Previous studies of other passerine birds have shown that high levels of parental effort may have an immunosuppressive effect. Our study indicates that weather conditions may override the effects of natural variation in parental effort, and that the PHA response is particularly influenced by short-term fluctuations in energy balance.     

Cell-mediated immunosenescence in birds

The phytohaemagglutinin (PHA) skin test response, used to assess cell-mediated immunity, is known to vary with many social and energetic factors, but the effects of age have received little attention. We found that the PHA response of immature birds was lower than those of the youngest breeding adults and were decreased in adults. Whenever possible, age should be included as a covariate when the PHA skin test is used to assess immunocompetence in ecological immunology. The rate of decline in PHA response differed between species and was inversely correlated with survival. The decrease in the PHA response averaged 57% over an average 80% of the maximum life span, but the absolute rate varied with species lifespan such that the short-lived species showed a greater loss per year than the long-lived species. This link between declining immune function and survival may reflect differences in resource partitioning between species, and suggests that selection may act on investment in immune function to influence maximum life span.     

不同繁殖状态雌性黑线仓鼠对植物血凝素的反应模式

由植物血凝素（Phytohemagglutinin,ＰＨＡ） 诱导产生的肿胀反应是生态免疫学中被广泛使用的免疫学参数,通过注射部位的肿胀程度大体上可反映细胞介导的免疫和天然免疫的综合变化情况。为探讨小型哺乳动物不同繁殖状态对ＰＨＡ 的反应模式,以非繁殖期、妊娠期、哺乳期和断乳期的雌性黑线仓鼠为研究对象,测定了注射ＰＨＡ 和生理盐水前（０ ｈ）及注射后６ ｈ、１２ ｈ 和２４ ｈ 注射部位皮肤组织的增厚程度。结果显示：（１）黑线仓鼠对ＰＨＡ 的反应有两种模式：非繁殖期、妊娠期和断乳期对ＰＨＡ 的反应模式相似,都在注射后６ ｈ 最高,１２ ｈ 和２４ ｈ 后逐渐下降;哺乳期对ＰＨＡ 的反应在注射后６ ｈ、１２ ｈ 和２４ ｈ 接近,彼此间的差异均不显著。（２）哺乳期黑线仓鼠的ＰＨＡ 反应高峰值有下降趋势,但与非繁殖期、妊娠期和断乳期高峰值之间的差异不显著。结果表明,黑线仓鼠在哺乳期对ＰＨＡ 的反应模式不同于其他繁殖阶段,且反应的高峰值被延迟,这对其存活可能有害,但有助于繁殖进程的延续和后代的生长发育。     

Severe respiratory viral infections: T-cell functions diverging from immunity to inflammation

Respiratory viral infections such as severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and influenza A virus (IAV) trigger distinct clinical outcomes defined by immunity-based viral clearance or disease associated with exaggerated and prolonged inflammation. The important role of T cells in shaping both antiviral immunity and inflammation has revived interest in understanding the host–pathogen interactions that lead to the diverse functions of T cells in respiratory viral infections. Inborn deficiencies and acquired insufficiency in immunity can prolong infection and shift the immune response towards exacerbated inflammation, which results from persistent innate immune activation and bystander T-cell activation that is nonspecific to the pathogen but is often driven by cytokines. This review discusses how virus variants, exposure doses, routes of infection, host genetics, and immune history can modulate the activation and function of T cells, thus influencing clinical outcomes. Knowledge of virus–host interaction can inform strategies to prevent immune dysfunction in respiratory viral infection and help in the treatment of associated diseases.     

Immunocompetence and helminth community of the white-toothed shrew, <Emphasis Type="Italic">Crocidura russula</Emphasis> from the Montseny Natural Park,Spain

Host immunocompetence assessed by spleen size and response to phytohaemagglutinin (PHA) injection may give some indications on the control of parasite infection and on host mediation effect, through immunity, on parasite community structure. We investigated the helminth community and immunocompetence of the white-toothed shrew in a small area to test the relationship between immunocompetence and intensity of helminth infection. At the proximate level and if spleen mass and PHA response reflect the level of immunocompetence, we expected that individuals with a large spleen or a high PHA response should harbour a lower parasite load than individuals with relatively small spleen or low PHA response. In addition, we predicted that the structure of the helminth community should be mediated by the host’s immune defence. Spleen mass was linked to helminth infection. Nematodes and cestodes were negatively associated within hosts. PHA response was not related to total helminth intensity of infection but was negatively related to cestode intensity and positively to nematode intensity. This result suggests either a differential modulating effect on immunity by the two groups of worms or the existence of an antagonistic association between nematodes and cestodes mediated by the immune response of the host.     

Maternal transfer of strain-specific immunity in an invertebrate

The most celebrated component of the vertebrate immune system is the acquired response in which memory cells established during primary infection enhance the proliferation of antibodies during secondary infection. Additionally, the strength of vertebrate acquired immune responses varies dramatically depending on the infecting pathogen species or on the pathogen genotype within species. Because invertebrates lack the T-cell receptors and Major Histocompatibility Complex (MHC) molecules that mediate vertebrate adaptive immune responses, they are thought to lack adaptive immunity and be relatively unspecific in their interactions with pathogens. With only innate immunity, invertebrate hosts are believed to be nai;ve at each new encounter with pathogens. Nevertheless, some forms of facultative immunity appear to be important in insects; some individuals have enhanced immunity due to population density, and some social insects benefit when their nest-mates have been exposed to a pathogen or pathogen mimic (; see for a predation example.) Here we provide evidence for acquired strain-specific immunity in the crustacean Daphnia magna infected with the pathogenic bacteria Pasteuria ramosa. Specifically, the fitness of hosts was enhanced when challenged with a bacterial strain their mother had experienced relative to cases when mother and offspring were challenged with different strains.     

No evidence for a trade-off between reproductive investment and immunity in a rodent

Life history theory assumes there are trade-offs between competing functions such as reproduction and immunity. Although well studied in birds, studies of the trade-offs between reproduction and immunity in small mammals are scarce. Here we examined whether reduced immunity is a consequence of reproductive effort in lactating Brandt's voles (Lasiopodomys brandtii). Specifically, we tested the effects of lactation on immune function (Experiment I). The results showed that food intake and resting metabolic rate (RMR) were higher in lactating voles (6≤ litter size ≤8) than that in non-reproductive voles. Contrary to our expectation, lactating voles also had higher levels of serum total Immunoglobulin G (IgG) and anti-keyhole limpet hemocyanin (KLH) IgG and no change in phytohemagglutinin (PHA) response and anti-KLH Immunoglobulin M (IgM) compared with non-reproductive voles, suggesting improved rather than reduced immune function. To further test the effect of differences in reproductive investment on immunity, we compared the responses between natural large (n≥8) and small litter size (n≤6) (Experiment II) and manipulated large (11-13) and small litter size (2-3) (Experiment III). During peak lactation, acquired immunity (PHA response, anti-KLH IgG and anti-KLH IgM) was not significantly different between voles raising large or small litters in both experiments, despite the measured difference in reproductive investment (greater litter size, litter mass, RMR and food intake in the voles raising larger litters). Total IgG was higher in voles with natural large litter size than those with natural small litter size, but decreased in the enlarged litter size group compared with control and reduced group. Our results showed that immune function is not suppressed to compensate the high energy demands during lactation in Brandt's voles and contrasting the situation in birds, is unlikely to be an important aspect mediating the trade-off between reproduction and survival.     

Cell-Mediated Immunity and Its Role in Resistance to Infection

The recently acquired knowledge of the importance of cell-mediated immunity in many illnesses and the discovery of a variety of substances that can restore certain cell-mediated immune functions has served to focus the attention of physicians on this area of immunity. It is important for practicing physicians to have a clear understanding of current knowledge of the role of cell-mediated immunity in resistance to infection and how this arm of the immune system relates to the diagnosis and therapy of infectious diseases.