植食性昆虫唾液效应子和激发子的研究进展

植食性昆虫与寄主植物通过协同进化形成了复杂的防御和反防御机制.本文系统综述了昆虫唾液效应子和激发子在植物与昆虫互作中的作用及机理.昆虫取食中释放的唾液激发子被植物识别而激活植物早期免疫反应,昆虫也能从口腔分泌效应子到植物体内抑制免疫;抗性植物则利用抗性(R)蛋白识别昆虫无毒效应子,启动效应子诱导的免疫反应,而昆虫又进化...     

Tsetse fly saliva biases the immune response to Th2 and induces anti-vector antibodies that are a useful tool for exposure assessment

Tsetse flies (Glossina sp.) are blood-feeding dipteran insects that transmit African trypanosomes, parasites that are responsible for human sleeping sickness and veterinary infections. Increasing attention is being paid to the effects of tsetse fly saliva deposited at the feeding site, which enables the blood-feeding process and putatively promotes parasite transmission. Here we demonstrate that saliva induces strong humoral responses against the major 43-45 kDa protein fraction (tsetse salivary gland proteins 1 and 2 - Tsal1 and Tsal2) in mice and humans and suppresses murine T and B cell responses to heterologous antigen. The saliva-induced immune response is associated with a Th2-biased cytokine profile and the production of mainly IgG1 and IgE antibody isotypes. Functionally, the antibodies raised in mice exposed to tsetse fly bites or induced after experimental saliva immunisation do not affect the fly's blood-feeding efficiency nor its survival. We propose that anti-saliva as well as anti-Tsal1/2 antibody responses can be used in epidemiological studies as a tool to analyze human exposure to tsetse flies.     

A functional genomics approach identifies candidate effectors from the aphid species Myzus persicae (green peach aphid)

Aphids are amongst the most devastating sap-feeding insects of plants. Like most plant parasites, aphids require intimate associations with their host plants to gain access to nutrients. Aphid feeding induces responses such as clogging of phloem sieve elements and callose formation, which are suppressed by unknown molecules, probably proteins, in aphid saliva. Therefore, it is likely that aphids, like plant pathogens, deliver proteins (effectors) inside their hosts to modulate host cell processes, suppress plant defenses, and promote infestation. We exploited publicly available aphid salivary gland expressed sequence tags (ESTs) to apply a functional genomics approach for identification of candidate effectors from Myzus persicae (green peach aphid), based on common features of plant pathogen effectors. A total of 48 effector candidates were identified, cloned, and subjected to transient overexpression in Nicotiana benthamiana to assay for elicitation of a phenotype, suppression of the Pathogen-Associated Molecular Pattern (PAMP)-mediated oxidative burst, and effects on aphid reproductive performance. We identified one candidate effector, Mp10, which specifically induced chlorosis and local cell death in N. benthamiana and conferred avirulence to recombinant Potato virus X (PVX) expressing Mp10, PVX-Mp10, in N. tabacum, indicating that this protein may trigger plant defenses. The ubiquitin-ligase associated protein SGT1 was required for the Mp10-mediated chlorosis response in N. benthamiana. Mp10 also suppressed the oxidative burst induced by flg22, but not by chitin. Aphid fecundity assays revealed that in planta overexpression of Mp10 and Mp42 reduced aphid fecundity, whereas another effector candidate, MpC002, enhanced aphid fecundity. Thus, these results suggest that, although Mp10 suppresses flg22-triggered immunity, it triggers a defense response, resulting in an overall decrease in aphid performance in the fecundity assays. Overall, we identified aphid salivary proteins that share features with plant pathogen effectors and therefore may function as aphid effectors by perturbing host cellular processes.     

The calcium activated nucleotidases: A diverse family of soluble and membrane associated nucleotide hydrolyzing enzymes

It has long been known that the salivary glands of hematophagous (blood-feeding) arthropods secrete soluble apyrases, which are potent nucleotide hydrolyzing enzymes capable of hydrolyzing extracellular ATP and ADP, the latter being a major agonist contributing to platelet aggregation. Only recently, however, has the identification of proteins homologous to these apyrases been reported in non-blood-feeding organisms such as rodents and humans. In this review, we present an overview of the diverse family of apyrases first described in the blood-feeding arthropods, including the identification and characterization of the soluble and membrane-bound vertebrate enzymes homologous to these arthropod apyrases. We also describe the enzymatic properties and nucleotide specificities of the expressed enzymes, and insights gained into the structure and function of this calcium activated nucleotidase (CAN) family from biophysical, mutagenesis and crystallography studies. The potential therapeutic value of these proteins is also discussed.     

唾液成分在刺吸式昆虫与植物关系中的作用

近年来,人们对刺吸式昆虫唾液成分的研究,揭示出其在刺吸式昆虫与植物关系中的重要作用。对多数刺吸式昆虫而言,他们取食时会分泌胶状和水状两种唾液,其中胶状唾液会在取食早期分泌形成唾液鞘来围绕并保护口针,通过直接和间接的作用来帮助取食;而水状唾液中则包含了果胶酶、纤维素酶、多酚氧化酶、过氧化物酶、碱性磷酸酯酶、蔗糖酶等组分,来帮助刺吸式昆虫对植物穿刺、消化食物、解毒次生物质并破坏植物的防御反应。有趣的是,唾液成分同时还可以诱导植物的防御反应,包括诱导植物的伤信号引起直接防御反应和诱导植物产生挥发物吸引植食者的天敌引起间接防御反应。并且,许多刺吸式昆虫取 食能够特异性地引发植物的病理反应,有研究推测刺吸式昆虫唾液中多聚半乳糖醛酸酶、碱性磷酸酯酶、蔗糖酶、多酚氧化酶等成分可能是某些植物特定病理反应的激发子,但是目前还没有定论,同时许多刺吸式昆虫唾液中的氨基酸和蛋白酶还是引起植物虫瘿的原因之一。 迄今的研究表明,刺吸式昆虫会根据不同的寄主植物和不同的生理需要,通过唾液组分的改变,来达到取食和发育的目的。对刺吸式昆虫唾液成分和作用机理的研究,可以为揭示刺吸式昆虫致害机理特别是传毒机理、指导害虫有效治理、阐明其与植物的协同进化等提供一定的思路。     

Quantitative proteomic analysis of the fall armyworm saliva

Lepidopteran larvae secrete saliva on plant tissues during feeding. Components in the saliva may aid in food digestion, whereas other components are recognized by plants as cues to elicit defense responses. Despite the ecological and economical importance of these plant-feeding insects, knowledge of their saliva composition is limited to a few species. In this study, we identified the salivary proteins of larvae of the fall armyworm (FAW), Spodoptera frugiperda; determined qualitative and quantitative differences in the salivary proteome of the two host races—corn and rice strains—of this insect; and identified changes in total protein concentration and relative protein abundance in the saliva of FAW larvae associated with different host plants. Quantitative proteomic analyses were performed using labeling with isobaric tags for relative and absolute quantification followed by liquid chromatography-tandem mass spectrometry. In total, 98 proteins were identified (>99% confidence) in the FAW saliva. These proteins were further categorized into five functional groups: proteins potentially involved in (1) plant defense regulation, (2) herbivore offense, (3) insect immunity, (4) detoxification, (5) digestion, and (6) other functions. Moreover, there were differences in the salivary proteome between the FAW strains that were identified by label-free proteomic analyses. Thirteen differentially identified proteins were present in each strain. There were also differences in the relative abundance of eleven salivary proteins between the two FAW host strains as well as differences within each strain associated with different diets. The total salivary protein concentration was also different for the two strains reared on different host plants. Based on these results, we conclude that the FAW saliva contains a complex mixture of proteins involved in different functions that are specific for each strain and its composition can change plastically in response to diet type.     

ATP hydrolyzing salivary enzymes of caterpillars suppress plant defenses

The oral secretions of herbivores are important recognition cues that can be used by plants to mediate induced defenses. In this study, a degradation of adenosine-5'-triphosphate (ATP) in tomato leaves was detected after treatment with Helicoverpa zea saliva. Correspondingly, a high level of ATPase activity in saliva was detected and three ATP hydrolyzing enzymes: apyrase, ATP synthase and ATPase 13A1 were identified in salivary glands. To determine the functions of these proteins in mediating defenses, they were cloned from H. zea and expressed in Escherichia coli. By applying the purified expressed apyrase, ATP synthase or ATPase 13A1 to wounded tomato leaves, it was determined that these ATP hydrolyzing enzymes suppressed the defensive genes regulated by the jasmonic acid and ethylene pathways in tomato plant. Suppression of glandular trichome production was also observed after treatment. Blood-feeding arthropods employ 5'-nucleotidase family of apyrases to circumvent host responses and the H. zea apyrase, is also a member of this family. The comparatively high degree of sequence similarity of the H. zea salivary apyrase with mosquito apyrases suggests a broader evolutionary role for salivary apyrases than previously envisioned.     

Effector proteins that modulate plant--insect interactions

Insect herbivores have highly diverse life cycles and feeding behaviors. They establish close interactions with their plant hosts and suppress plant defenses. Chewing herbivores evoke characteristic defense responses distinguishable from general mechanical damage. In addition, piercing-sucking hemipteran insects display typical feeding behavior that suggests active suppression of plant defense responses. Effectors that modulate plant defenses have been identified in the saliva of these insects. Tools for high-throughput effector identification and functional characterization have been developed. In addition, in some insect species it is possible to silence gene expression by RNAi. Together, this technological progress has enabled the identification of insect herbivore effectors and their targets that will lead to the development of novel strategies for pest resistances in plants.     

Evolution of the salivary apyrases of blood-feeding arthropods

Phylogenetic analyses of three families of arthropod apyrases were used to reconstruct the evolutionary relationships of salivary-expressed apyrases, which have an anti-coagulant function in blood-feeding arthropods. Members of the 5′nucleotidase family were recruited for salivary expression in blood-feeding species at least five separate times in the history of arthropods, while members of the Cimex-type apyrase family have been recruited at least twice. In spite of these independent events of recruitment for salivary function, neither of these families showed evidence of convergent amino acid sequence evolution in salivary-expressed members. On the contrary, in the 5′-nucleotide family, salivary-expressed proteins conserved ancestral amino acid residues to a significantly greater extent than related proteins without salivary function, implying parallel evolution by conservation of ancestral characters. This unusual pattern of sequence evolution suggests the hypothesis that purifying selection favoring conservation of ancestral residues is particularly strong in salivary-expressed members of the 5′-nucleotidase family of arthropods because of constraints arising from expression within the vertebrate host.     

Secretory proteins from the salivary glands of adult Aedes aegypti mosquitoes

We determined whether the salivary proteins of mosquitoes are constant throughout adult life and whether the nature of the feeding stimulus determined salivary composition. Acrylamide gel electrophoresis and subsequent silver-staining of harvested saliva yielded as many as 20 polypeptides; those precipitating silver most readily generally incorporated abundant isotope. Autoradiography revealed 12 prominant secretory polypeptide fractions, each synthesized beginning soon after ecdysis and throughout the first week of adult life. Synthesis was limited to the posterior regions of the salivary gland, mainly within the lateral lobes. Virtually all secretory polypeptides were antigenic and were secreted into hosts during blood-feeding. We concluded that the secretory salivary proteins of adult mosquitoes are continuously synthesized throughout the first week of adult life, and that most are secreted during blood-feeding.     

Shaping the understanding of saliva-derived effectors towards aphid colony proliferation in host plant

‘Effectors’ are proteins and/or small molecules that originated from aphid saliva gland and its secretion is initiated due to interaction between host and insect. The effectors have the ability to manipulate the host cell structure as well as function similar to pathogen’s effectors. Like pathogen’s effectors, aphid effectors suppress the hosts’ defense responses as well as hosts’ defense induction or both. In the susceptible interaction with the host, aphid effectors alter plant processes that contribute to the establishment of compatibility that promotes aphid proliferation. In the susceptible reaction with the host, aphid effectors contribute to the successful salivation and sustainability of the sieve element sap ingestion that have promoting role in more aphid proliferation. In the resistant interaction with the host, aphid effectors are recognized by the typical plant receptors and elicit the induction the effective defense response. As a result, aphid proliferation is reduced due to reduced compatibility establishment in the resistant host. This review focuses on the exciting progress in aphid effector biology that insights new perspective in the molecular basis of plant–aphid interactions.     

Probing the importance of hydrogen bonds in the active site of the subtilisin nattokinase by site-directed mutagenesis and molecular dynamics simulation

Apyrase activity is present in the saliva of haematophagous arthropods. It is related to blood-feeding because of the apyrase ability to hydrolyse ADP, a key component of platelet aggregation. Five apyrases with apparent molecular masses of 88, 82, 79, 68 and 67 kDa were identified in the saliva of the vector of Chagas disease, Triatoma infestans. The large size observed during purification of these enzymes suggested oligomerization. In the present study, we confirmed, using gel-filtration and analytical ultracentrifugation, the presence of apyrase oligomers with molecular masses of 200 kDa in the saliva. Electrophoretic analyses showed that disulphide bonds were involved in homo-oligomerization. In addition, heterogeneity in disulphide bonds and in pI was detected, with the pI ranging from 4.9 to 5.4. The present study gives the first insights into the quaternary structure of soluble apyrases.     

Cooperative Blood-feeding and the Function and Implications of Feeding Aggregations in the Sand Fly,Lutzomyia longipalpis (Diptera: Psychodidae)

Given the importance that the evolution of cooperation bears in evolutionary biology and the social sciences, extensive theoretical work has focused on identifying conditions that promote cooperation among individuals. In insects, cooperative or altruistic interactions typically occur amongst social insects and are thus explained by kin selection. Here we provide evidence that in Lutzomia longipalpis, a small biting fly and an important vector of leishmaniasis in the New World, cooperative blood-feeding in groups of non-kin individuals results in a strong decrease in saliva expenditure. Feeding in groups also strongly affected the time taken to initiate a bloodmeal and its duration and ultimately resulted in greater fecundity. The benefits of feeding aggregations were particularly strong when flies fed on older hosts pre-exposed to sand fly bites, suggesting that flies feeding in groups may be better able to overcome their stronger immune response. These results demonstrate that, in L. longipalpis, feeding cooperatively maximizes the effects of salivary components injected into hosts to facilitate blood intake and to counteract the host immune defences. As a result, cooperating sand flies enjoy enormous fitness gains. This constitutes, to our knowledge, the first functional explanation for feeding aggregations in this species and potentially in other hematophagous insects and a rare example of cooperation amongst individuals of a non-social insects species. The evolution of cooperative group feeding in sand flies may have important implications for the epidemiology of leishmaniasis.     

Leishmania parasites: could we consider them as living organisms per se?

Over the last 10 years - in Microbes and Infection - the publications dealing with protozoan parasites were mainly providing insights on the pathogenic processes leading to the local or systemic damages in the mammals, these parasitic organisms exploit/subvert as hosts. As a result, many investigators introduced the objectives of their analysis by referring to "host-pathogen" interactions. Though we, as investigators, are all determined to decipher the pathogenic processes which can indeed be coupled to the parasite uncontrolled development, I think that the parasites - alike the living organisms they subvert as hosts - need to be considered as living organisms per se, instead of being considered as "pathogens". Such a conceptual frame will promote research on the processes on which relies their perpetuation whatever the level under investigations - individual and/or population level. Only the unicellular protozoan parasites of the genus Leishmania known to be hosted by blood-feeding insects and mammals will be further considered in this brief contribution.     

The biological and immunomodulatory properties of sand fly saliva and its role in the establishment of Leishmania infections

Sand fly saliva contains a rich array of pharmacologically active compounds whose primary function is to prevent the hemostatic mechanisms of the host. Several studies have ascribed immunosuppressive properties to sand fly saliva as well as an exacerbative effect on Leishmania infectivity for their mammalian hosts. This review provides a comprehensive account of sand fly salivary components, the immunomodulatory properties exhibited by some of its molecules, and describes the findings concerning the influence of saliva on Leishmania infections. The potential use of saliva as part of an anti-Leishmania vaccine for the mammalian host is also addressed.     

Differential modulation of murine host immune response by salivary gland extracts from the mosquitoes Aedes aegypti and Culex quinquefasciatus

Mosquitoes (Diptera: Culicidae) are major vectors of numerous infectious agents. Compounds in mosquito saliva not only facilitate blood-feeding, but may also have an impact upon the immune system of vertebrate hosts. Consequently, the exposure to mosquito saliva may influence pathogen transmission, establishment and disease development. Using two medically important vector mosquitoes, Aedes aegypti (L.) and Culex quinquefasciatus Say, we examined the effects of mosquito saliva on immune cells of host mice. After antigen-specific or non-specific stimulation, murine splenocyte proliferation and production of both Th1 and Th2 cytokines were significantly reduced in the presence of salivary gland extract (SGE) from Ae. aegypti, but not SGE from Cx. quinquefasciatus. T cell populations were highly susceptible to this suppression, showing increased mortality and reduced division rates - judged by flow cytometric analyses. Evidently these two culicine mosquitoes differ in their host immunomodulatory activities.