Testing the effect of individual color morphology on immune response in bush-crickets

Abstract Despite the growing interest in how an individual's immune response is correlated to morphological and ecological factors, little empirical data has been available from wild insect populations. Many insects have different color morphs and exhibit differences in immune responses. Links are expected to exist between body colors and immune function in insects, because the same biochemical precursors involved in the immune response are responsible for melanin-based color markings. In this study, I assay the immune response of two different color morphs of 377 wild-caught bush-crickets, Metrioptera roeseli, from 20 populations by measuring individual encapsulation responses to a surgically implanted nylon monofilament. There was no difference between green and brown bush-cricket morphs (low melanin vs high melanin investment in cuticula color respectively) and their ability to mount an immune response to the implant. Further study is needed on the relationship between color morphology and immune response in wild insects, and whether trade-offs occur between factors during an insect's development phase and long-term health.     

Immune response to nylon filaments in two damselfly species that differ in their resistance to ectoparasitic mites

1. Insects commonly resist parasites using melanotic encapsulation. Many studies measuring immune response use the amount of melanin deposited on an artificial object that has been inserted into the animal as a proxy of the amount of resistance that the host is capable of mounting to natural parasites. 2. The relevance of this methodology to immune response in natural insect populations needs further study. Here, we examined two temperate damselfly species to elucidate the relationships among damselfly size, natural resistance to mites, and the immune response mounted by the same damselflies against nylon filaments. 3. The damselfly species that had high rates of melanotic encapsulation of mites in nature deposited more melanin on the nylon inserts than the species with low rates of natural resistance. 4. In females of this species, those that had resisted mites naturally melanised the nylon filament more aggressively than those that did not resist mites. 5. Our results show some support for the use of nylon filaments to assess natural patterns of immune response in these damselflies, but also suggest that caution should be used in interpreting the responses.     

Synergistic and antagonistic interaction between different branches of the immune system is related to melanin‐based coloration in nestling tawny owls

When exposed to parasites, hosts often mount energetically expensive immune responses, and this may alter resource allocation between competing life history traits including other components of the immune system. Here, we investigated whether a humoral immune challenge towards a vaccine reduces or enhances the cutaneous immune responses towards an injection of lipopolysaccharid (LPS, innate immunity) and phytohaemagglutinin (PHA, T‐cell immunity) in nestling tawny owls in interaction with the degree of plumage melanin‐based coloration. The humoral immune challenge enhanced the response to LPS similarly in differently coloured nestlings. In contrast, the same humoral immune challenge enhanced immune response to PHA in dark reddish melanic nestlings while reducing it in pale reddish melanic nestlings. Our results highlight that both antagonistic and synergistic interactions can take place among branches of immune system, and that the sign and magnitude of these interactions can vary with immune responses involved and the degree of melanin‐based coloration.     

Morphofunctional characterization of hemocytes in black soldier fly larvae

In insects, the cell-mediated immune response involves an active role of hemocytes in phagocytosis, nodulation, and encapsulation. Although these processes have been well documented in multiple species belonging to different insect orders, information concerning the immune response, particularly the hemocyte types and their specific function in the black soldier fly Hermetia illucens, is still limited. This is a serious gap in knowledge given the high economic relevance of H. illucens larvae in waste management strategies and considering that the saprophagous feeding habits of this dipteran species have likely shaped its immune system to efficiently respond to infections. The present study represents the first detailed characterization of black soldier fly hemocytes and provides new insights into the cell-mediated immune response of this insect. In particular, in addition to prohemocytes, we identified five hemocyte types that mount the immune response in the larva, and analyzed their behavior, role, and morphofunctional changes in response to bacterial infection and injection of chromatographic beads. Our results demonstrate that the circulating phagocytes in black soldier fly larvae are plasmatocytes. These cells also take part in nodulation and encapsulation with granulocytes and lamellocyte-like cells, developing a starting core for nodule/capsule formation to remove/encapsulate large bacterial aggregates/pathogens from the hemolymph, respectively. These processes are supported by the release of melanin precursors from crystal cells and likely by mobilizing nutrient reserves in newly circulating adipohemocytes, which could thus trophically support other hemocytes during the immune response. Finally, the regulation of the cell-mediated immune response by eicosanoids was investigated.     

Melanin‐based coloration is related to parasite intensity and cellular immune response in an urban free living bird: the feral pigeon Columba livia

Melanin‐based coloration is widespread among vertebrates, but the adaptive function of this trait remains poorly known. Recently, it has been shown that differently coloured individuals have different abilities to cope with parasites. This correlation between melanin‐based coloration and immunity could be explained by the pleiotropic effects of genes coding for melanin pigmentation on the immune system (‘genetic link’ hypothesis) but also because differently coloured individuals may exploit alternative habitats varying in parasite exposure, which leads to different development of the immune function (‘exposure’ hypothesis). As feral pigeons Columba livia are genetically polymorphic with respect to melanic coloration, they constitute an ideal model system to address such hypotheses. In this study, we showed that darker melanic individuals had a lower endoparasite intensity (reflecting host susceptibility) and had a greater cellular immune response to PHA injection than paler ones, whereas parasite prevalence (reflecting exposure to vectors) was similar between colorations. These results provide a correlative support of the ‘genetic link’ hypothesis: differently coloured individuals might be similarly exposed to parasites but darker ones might have a better ability to control the infection. This suggests that parasitism could play a crucial role in the maintenance of colour polymorphism in natural populations, which opens the interesting possibility that differently coloured individuals could be adapted to alternative environments varying in parasite diversity and exposure.     

Sedimentation rapidly induces an immune response and depletes energy stores in a hard coral

High sedimentation rates have been linked to reduced coral health within multiple systems; however, whether this is a direct result of compromised coral immunity has not been previously investigated. The potential effects of sedimentation on immunity of the hard coral Montipora patula were examined by comparing physiological responses of coral fragments inoculated with sterilized marine sediments and those under control conditions. Sediments were collected from terrestrial runoff-affected reefs in SW Madagascar and applied cyclically for a total of 24 h at a rate observed during precipitation-induced sedimentation events. Coral health was determined 24 h after the onset of the sedimentation stress through measuring metabolic proxies of O₂ budget and lipid ratios. Immune response of the melanin synthesis pathway was measured by quantifying phenoloxidase activity and melanin deposits. Sedimentation induced both immune and metabolic responses in M. patula. Both phenoloxidase activity and melanin deposition were significantly higher in the sediment treatment compared to controls, indicating an induced immune response. Sediment-treated corals also showed a tendency towards increased respiration (during the night) and decreased photosynthesis (during the day) and a significant depletion of energy reserves as compared to controls. These data highlight that short-term (24 h) sedimentation, free of live microorganisms, compromises the health of M. patula. The energetically costly immune response, potentially elicited by residual endotoxins and other inflammatory particles associated with the sterile sediments, likely contributes to the energy depletion. Overall, exposure to sedimentation adversely affects coral health and continued exposure may lead to resource depletion and an increased susceptibility to disease.     

Ultrastructure of the melanization response of Aedes trivittatus against inoculated Dirofilaria immitis microfilariae

Scanning and transmission electron microscopy revealed that intrathoracically-inoculated microfilariae (mff) of Dirofilaria immitis elicited a rapid and effective immune response in the hemocoel of Aedes trivittatus mosquitoes. Hemocyte lysis and melanization of inoculated mff began immediately following exposure to the hemolymph environment. Initial melanin accumulation occurred at any site along the surface of mff and rapidly increased in thickness. Hemocyte encapsulation generally described for insects did not occur, but hemocytes might be necessary for activation of the melanization response. Although intact hemocytes were never abundant, those that were present seemed to show an active secretion of membrane-bound vacuoles directed toward mff. Activated hemocytes were in close association, but never in direct contact with the parasite, and were most commonly seen in various stages of lysis. Numerous cell remnants were noted throughout the developing melanin capsule. Parasites were completely melanized by 24 hr postinoculation (PI). By about 3 days PI, a membrane began to form around deposited melanin and hemocyte remnants. This developed into a double membrane-like structure of 25-30 nm thickness and resulted in the enclosure and isolation of the mff, melanin deposits, and cellular remnants from hemolymph components. It is suggested that this membrane functions as a boundary to isolate the melanized parasite and prevents additional hemocyte involvement.     

Multivariate heredity of melanin-based coloration,body mass and immunity

The genetic covariation among different traits may cause the appearance of correlated response to selection on multivariate phenotypes. Genes responsible for the expression of melanin-based color traits are also involved in other important physiological functions such as immunity and metabolism by pleiotropy, suggesting the possibility of multivariate evolution. However, little is known about the relationship between melanin coloration and these functions at the additive genetic level in wild vertebrates. From a multivariate perspective, we simultaneously explored inheritance and selection of melanin coloration, body mass and phytohemagglutinin (PHA)-mediated immune response by using long-term data over an 18-year period collected in a wild population of the common kestrel Falco tinnunculus. Pedigree-based quantitative genetic analyses showed negative genetic covariance between melanin-based coloration and body mass in male adults and positive genetic covariance between body mass and PHA-mediated immune response in fledglings as predicted by pleiotropic effects of melanocortin receptor activity. Multiple selection analyses showed an increased fitness in male adults with intermediate phenotypic values for melanin color and body mass. In male fledglings, there was evidence for a disruptive selection on rump gray color, but a stabilizing selection on PHA-mediated immune response. Our results provide an insight into the evolution of multivariate traits genetically related with melanin-based coloration. The differences in multivariate inheritance and selection between male and female kestrels might have resulted in sexual dimorphism in size and color. When pleiotropic effects are present, coloration can evolve through a complex pathway involving correlated response to selection on multivariate traits.     

Darker female pigeons transmit more specific antibodies to their eggs than do paler ones

Melanin‐based coloration is widespread among vertebrates, yet the adaptive significance of such pigments remains elusive, particularly with regard to the link between melanin and immune‐mediated maternal effects. The aim of this study was to investigate whether melanin‐based coloration could signal the ability of mothers to mount a humoral response and to transfer maternal antibodies (Ab) to their young. We injected differently coloured (pale and dark) female feral pigeons (Columba livia) with Chlamydiae (a natural antigen) and Keyhole Limpet Haemocyanin (KLH, an artificial antigen), and found no significant difference in humoral response between differently coloured females. However, darker females transferred more Ab against Chlamydiae into their eggs than paler ones, despite similar circulating levels of Ab. In addition to this, melanin‐based coloration showed a high heritability value. This suggests that a genetically based coloured trait might be linked to the ability of females to transfer specific Ab against Chlamydiae (but not against KLH) to their offspring, independent of their ability to produce Ab. This suggests that transmission of maternal Ab is antigen dependent, and that melanin‐based coloration might signal female ability to transmit specific Ab against natural pathogens. © 2013 The Linnean Society of London     

Fluorescent quantification of melanin

Melanin quantification is reportedly performed by absorption spectroscopy, commonly at 405 nm. Here, we propose the implementation of fluorescence spectroscopy for melanin assessment. In a typical in vitro assay to assess melanin production in response to an external stimulus, absorption spectroscopy clearly overvalues melanin content. This method is also incapable of distinguishing non‐melanotic/amelanotic control cells from those that are actually capable of performing melanogenesis. Therefore, fluorescence spectroscopy is the best method for melanin quantification as it proved to be highly specific and accurate, detecting even small variations in the synthesis of melanin. This method can also be applied to the quantification of melanin in more complex biological matrices like zebrafish embryos and human hair.     

黑色素羽毛装饰反映了鸟类的抗氧化和免疫能力吗?

鸟类信号系统研究认为,不同于类胡萝卜素,基于黑色素的羽色装饰不需要昂贵的生理代价。然而,羽毛色素细胞的黑色素沉积与抗氧化能力和免疫系统有很强的联系,而抗氧化能力和免疫系统在提高有机体适合度方面具有重要的功能。黑色素细胞的生长对氧化环境压力十分敏感;并且,黑色素本身似乎就具有抗氧化剂的功能。相应地,把抗氧化剂用于以黑色素为基础羽色发育,还是用于其它方面例如免疫调节和免疫刺激等,个体也许必须对此做出权衡。组织中的抗氧化功能大多与代谢活动有关,也就是和自由基的最高水平有关。此外,人们发现,在哺乳动物中调节黑色素沉积的激素,即α黑素细胞刺激素,在鸟类上皮组织的色素沉积中也具有同样的功能。这种进化保守的激素是免疫和炎症反应的一个重要介体。它下调前炎性细胞因子、免疫介导细胞因子和协同刺激分子,以及主要组织相容性复合体Ⅰ类分子在单核细胞的表达以及抗体的产生,而上调抑制因子白介素。黑色素在羽毛上的大量沉积,也可能反映出免疫系统在局部免疫应答的负调控能力。这些将黑色素沉积和抗氧化剂以及免疫功能联系起来的机制,表明基于黑色素的羽色信号具有一定的生理代价,这些对以往关于黑色素的优势作用的假说提出质疑     

A novel 43-kDa protein as a negative regulatory component of phenoloxidase-induced melanin synthesis

The melanization reaction induced by activated phenoloxidase in arthropods is important in the multiple host defense innate immune reactions, leading to the sequestration and killing of invading microorganisms. This reaction ought to be tightly controlled because excessive formation of quinones and systemic hypermelanization are deleterious to the hosts, suggesting that a negative regulator(s) of melanin synthesis may exist in hemolymph. Here, we report the purification and cloning of a cDNA of a novel 43-kDa protein, from the meal-worm Tenebrio molitor, which functions as a melanization-inhibiting protein (MIP). The deduced amino acid sequence of 352 residues has no homology to known sequences in protein data bases. When the concentration of the 43-kDa protein was examined by Western blot analysis in a melanin-induced hemolymph prepared by injection of Candida albicans into T. molitor larvae, the 43-kDa protein specifically decreased in the melanin-induced hemolymph compared with control hemolymph. Recombinant MIP expressed in a baculovirus system had an inhibitory effect on melanin synthesis in vitro. RNA interference using a synthetic 445-mer double-stranded RNA of MIP injected into Tenebrio larvae showed that melanin synthesis was markedly induced. These results suggest that this 43-kDa MIP inhibits the formation of melanin and thus is a modulator of the melanization reaction to prevent the insect from excessive melanin synthesis in places where it should be inappropriate.     

Dorsal pattern polymorphism in female Iberian wall lizards: differences in morphology,dorsal coloration,immune response,and reproductive investment

Sex‐specific colour polymorphisms have been extensively documented in many different taxa. When polymorphism in colour pattern is restricted to females, the condition is known as female‐limited pattern polymorphism (FPP), which has been less commonly addressed in vertebrates. FPP is present in several lizard species, although most research on lizards has focused on carotenoid‐ and pteridine‐based coloration and not on melanin‐based polymorphisms. In the present study, we focus on Iberian wall lizards, Podarcis hispanicus, where two female melanin‐based dorsal patterns can be clearly distinguished: striped and reticulated‐blotched. We indirectly tested the hypothesis that selection acts differentially among P. hispanicus female morphs to create alternative morph‐specific phenotypic optima at different levels by investigating whether morphs differ in fitness proxies. We specifically examined whether the two female dorsal pattern morphs differed in adult morphology, dorsal coloration, immune response, reproductive investment, and growth. We did not find a relationship between melanin‐based coloration and hatchling growth and immune response, despite a correlation between these traits possibly being expected as a result of pleiotropy in the melanocortin system. However, our results show that female dorsal morphs in P. hispanicus differ in terms of adult morphology, dorsal coloration, and reproductive investment. Reticulated‐blotched P. hispanicus females had deeper heads and longer femora, less melanin, and more brownish coloration, and also had larger and heavier hatchlings than striped females.     

Immune function responds to selection for cuticular colour in Tenebrio molitor

Cuticular colour in the mealworm beetle (Tenebrio molitor) is a quantitative trait, varying from tan to black. Population level variation in cuticular colour has been linked to pathogen resistance in this species and in several other insects: darker individuals are more resistant to pathogens. Given that cuticular colour has a heritable component, we have taken an experimental evolution approach: we selected 10 lines for black and 10 lines for tan adult cuticular phenotypes over at least six generations and measured the correlated responses to selection in a range of immune effector systems. Our results show that two immune parameters related to resistance (haemocyte density and pre-immune challenge activity of phenoloxidase (PO)) were significantly higher in selection lines of black beetles compared to tan lines. This may help to explain increased resistance to pathogens in darker individuals. Cuticular colour is dependent upon melanin production, which requires the enzyme PO that is present in its inactive form inside haemocytes. Thus, the observed correlated response to selection upon cuticular colour and immune variables probably results from these traits' shared dependence on melanin production.     

The mathematics of tanning

Background  

The pigment melanin is produced by specialized cells, called melanocytes. In healthy skin, melanocytes are sparsely spread among the other cell types in the basal layer of the epidermis. Sun tanning results from an UV-induced increase in the release of melanin to neighbouring keratinocytes, the major cell type component of the epidermis as well as redistribution of melanin among these cells. Here we provide a mathematical conceptualization of our current knowledge of the tanning response, in terms of a dynamic model. The resolution level of the model is tuned to available data, and its primary focus is to describe the tanning response following UV exposure.     

Cuticular melanisation and immune response in a butterfly: local adaptation and lack of correlation

1. Despite growing knowledge on the relationship between ecological variables and individual immune function, data on the spatial variability of immune defence in invertebrate natural populations are scarce. 2. Here, we use replicated populations of the butterfly Lycaena tityrus from different altitudes to investigate genetic variation in the melanin‐based encapsulation response. As high‐ and low‐altitude populations differ in cuticular pupal melanisation, we further tested for any associations between pupal melanisation and parasite resistance. 3. Although pupal melanisation was higher at higher compared with lower altitudes (and at a higher compared with a lower rearing temperature), any obvious relations to the encapsulation response were absent. Further phenotypic correlations within groups were significant in one out of four cases only, suggesting that in L. tityrus encapsulation operates largely independent of cuticular melanisation. 4. A significant interaction between altitude and temperature indicated that high‐altitude animals show a stronger melanisation response than low‐altitude ones at the lower temperature and vice versa, indicating local adaptation to different climates.     

A mechanistic link between parasite resistance and expression of a sexually selected trait in a damselfly

This paper examines a field-based insect system in which a signal trait and an immune effector system responsible for parasite resistance rely on the same melanin-producing enzyme cascade (phenoloxidase, PO). Observations and experiments on males of the calopterygid damselfly Calopteryx splendens xanthostoma revealed that resistance to the prevalent parasite in the study system (a eugregarine protozoan infecting the mid-gut) was correlated with quantitative aspects of the sexually dimorphic melanized wingspot of males, a trait that is produced and fixed before the host comes into contact with the sporozsoites of the parasite. Regulation of PO during experimental immune challenge showed that males with dark, homogenous melanin distribution in their wings showed no change in PO levels 24 h after challenge. By contrast males with lighter and/or more heterogenous melanin distribution in their wings tended to show higher PO levels 2-h after immune challenge. The changes in PO levels occur despite the lack of a relationship between wing-pigment distribution and the cellular encapsulation response. These results suggest a shared, limiting resource may form the mechanistic basis of the trade-off between a condition-dependent signal trait and immune function in this system.     

Induction of immune tolerance by dendritic cells: implications for preventative and therapeutic immunotherapy of autoimmune disease

Dendritic cells (DC) have a key role in controlling the immune response, by determining the outcome of antigen presentation to T cells. Through costimulatory molecules and other factors, DC are involved in the maintenance of peripheral tolerance through modulation of the immune response. This modulation occurs both constitutively, and in inflammation, in order to prevent autoimmunity and to control established immune responses. Dendritic cell control of immune responses may be mediated through cytokine or cell-contact dependent mechanisms. The molecular and cellular basis of these controls is being understood at an increasingly more complex level. This understanding is reaching a level at which DC-based therapies for the induction of immune regulation in autoimmunity can be tested in vivo. This review outlines the current state of knowledge of DC in immune tolerance, and proposes how DC might control both T cell responses, and themselves, to prevent autoimmunity and maintain peripheral tolerance.