Separation and behavior in vitro of hemocytes from the moth,Pseudoplusia includens

Hemocytes collected from larvae of Pseudoplusia includens (Lepidoptera: Noctuidae) were separated by centrifugation on Percoll cushions. The procedure resulted in 95% purity of plasmatocytes and greater than 99% purity of granular and spherule cells. Medium supplemented with chicken serum enhanced cell viability and promoted spreading of plasmatocytes. Cell-free plasma and medium preconditioned by plasmatocytes or granular cells stabilized cells in vitro and also accelerated spreading of plasmatocytes relative to medium supplemented with chicken serum. Oenocytoids were the only morphotype that exhibited endogenous phenoloxidase activity, while granular cells and plasmatocytes were the only cells that endocytosed fluorescent beads in vitro. Granular cells and plasmatocytes ingested fluorescently labelled beads, both in mixed populations of hemocytes and after separation. Plasmatocytes were the only morphotype that encapsulated large foreign targets in vitro following separation. Separated granular cells attached and spread on the surface of foreign targets but never formed a multilayered capsule.     

Monoclonal antibody MS13 identifies a plasmatocyte membrane protein and inhibits encapsulation and spreading reactions of Manduca sexta hemocytes

Lepidopterans generally can successfully defend themselves against a variety of parasites or parasitoids. One mechanism they use is to encapsulate the invader in many layers of hemocytes. For encapsulation to occur, the hemocytes must attach to the foreign material, spread, and adhere to each other. The molecules that mediate these processes are not known. One method to identify proteins potentially necessary for adhesion, spreading, and, thus, encapsulation is to use monoclonal antibodies that interfere with these functions. In this paper, we report that a monoclonal antibody against Manduca sexta plasmatocytes effectively inhibited encapsulation of synthetic beads in vitro and in vivo. Furthermore, it inhibited plasmatocyte spreading in vitro. Other anti-hemocyte antibodies did not have these effects. The plasmatocyte-specific monoclonal antibody, mAb MS13, recognized a protein of approximately 90,000 daltons as indicated by Western blot analysis of hemocyte lysate proteins. The epitope recognized by mAb MS13 was present on the exterior surface of plasmatocytes. Using indirect immunohistochemistry with hemocyte-specific antibodies, we also determined that during encapsulation plasmatocytes were the first cells bound to latex beads and later layers consisted of both plasmatocytes and granular cells. Arch.     

Microplitis demolitor bracovirus inhibits phagocytosis by hemocytes from Pseudoplusia includens

The braconid wasp Microplitis demolitor carries Microplitis demolitor bracovirus (MdBV) and parasitizes the larval stage of several noctuid moths. A key function of MdBV in parasitism is suppression of the host's cellular immune response. Prior studies in the host Pseudoplusia includens indicated that MdBV blocks encapsulation by preventing two types of hemocytes, plasmatocytes and granulocytes, from adhering to foreign targets. The other main immune response mediated by insect hemocytes is phagocytosis. The goal of this study was to determine which hemocyte types were phagocytic in P. includens and to assess whether MdBV infection affects this defense response. Using the bacterium Escherichia coli and inert polystyrene beads as targets, our results indicated that the professional phagocyte in P. includens is granulocytes. The phagocytic responses of granulocytes were very similar to those of High Five cells that prior studies have suggested are a granulocyte-like cell line. MdBV infection dose-dependently disrupted phagocytosis in both cell types by inhibiting adhesion of targets to the cell surface. The MdBV glc1.8 gene encodes a cell surface glycoprotein that had previously been implicated in disruption of adhesion and encapsulation responses by immune cells. Knockdown of glc1.8 expression by RNA interference (RNAi) during the current study rescued the ability of MdBV-infected High Five cells to phagocytize targets. Collectively, these results indicate that glc1.8 is a key virulence determinant in disruption of both adhesion and phagocytosis by insect immune cells.     

Ex vivo observation of insect hemocyte behavior against beads and nematodes in the presence of insect plasma

Recognition of foreign targets by insect hemocytes is a crucial first step for insect immunity against invading multicellular organisms in the hemocoel. To understand the mechanism of recognition, we observed the hemocyte behavior of Galleria mellonella Linnaeus (Lepidoptera: Pyralidae) larvae against beads and the nonparasitic nematode Caenorhabditis elegans (Maupas) (Rhabditida: Rhabditidae) in the presence of plasma ex vivo using time-lapse microscopy. Both granular cells and plasmatocytes adhered to and spread on the surface of beads and nematodes. In addition, the spread plasmatocytes actively moved over the beads and nematodes. These results suggest that not only granular cells but also plasmatocytes can recognize foreign targets in the presence of insect plasma and that spread plasmatocytes can actively search for foreign targets. Hemocyte adhesion to beads and nematodes ex vivo was similar to that of the in vivo 1?h after injection. A divalent cation chelator inhibited the spreading and adhesion of plasmatocytes ex vivo, but it did not affect the adhesion of granular cells. The present method enables the analysis of acute hemocyte response against foreign targets in the presence of plasma.

     

Plasmatocyte sensitivity to plasmatocyte spreading peptide (PSP) fluctuates with the larval molting cycle

Plasmatocyte spreading peptide (PSP) is a cytokine from the moth Pseudoplusia includens that activates a class of hemocytes called plasmatocytes to bind and spread on foreign surfaces. Previous structure-function studies on PSP used plasmatocytes collected from P. includens larvae that were in the late stages of the last (fifth) instar. Here, we report that plasmatocyte sensitivity to PSP varied significantly during the fourth and fifth instar. PSP weakly activated plasmatocytes early in the instar when hemolymph juvenile hormone (JH) titers were relatively high and ecdysteroid titers were low, but strongly activated plasmatocytes late in the instar after JH titers declined and ecdysteroid titers rose. In contrast, plasmatocytes did not vary in their response to plasma, which contains other factors besides PSP that affect plasmatocyte function. In vitro assays indicated that 20-hydroxyecdysone (20E) dose-dependently synergized PSP activity, whereas the JH analog methoprene antagonized PSP activity. Methoprene had no effect on adhesion and spreading of granular cells, but plasmatocytes from larvae topically treated with methoprene exhibited a reduction in sensitivity to PSP. Collectively, these results indicate that plasmatocyte sensitivity to PSP fluctuates in relation to the molting cycle, and that PSP activity is affected by juvenoids and ecdysone.     

Plasmatocytes from the moth Pseudoplusia includens induce apoptosis of granular cells

The primary immune response toward internal parasites and other large foreign objects that enter the insect hemocoel is encapsulation. Prior studies indicated that granular cells and plasmatocytes are the two hemocyte types required for capsule formation by the moth Pseudoplusia includens (Lepidoptera: Noctuidae). Capsules formed by P. includens also have a defined architecture with primarily granular cells attaching directly to the target, multiple layers of plasmatocytes adhering to this inner layer of granular cells, and a monolayer of granular cells attaching to the capsule periphery. Dye-exclusion assays indicated that granular cells die shortly after attaching to the capsule periphery, leaving a basal lamina-like layer around the capsule. In examining the mechanisms underlying granular cell death, we found that culture medium preconditioned by plasmatocytes induced apoptosis of granular cells. Characteristics of plasmatocyte-induced apoptosis included condensation of chromatin, cell surface blebbing and fragmentation of nuclear DNA. Plasmatocyte-conditioned medium did not induce apoptosis of other hemocyte types, and medium conditioned by other hemocyte types did not induce apoptosis of granular cells. The adhesive state of granular cells and plasmatocytes also affected levels of apoptosis. Conditioned medium from spread plasmatocytes induced higher levels of granular cell apoptosis than medium conditioned by unspread plasmatocytes. Reciprocally, spread granular cells underwent significantly higher rates of apoptosis than unspread granular cells in medium conditioned by spread plasmatocytes. In situ analysis indicated that granular cells on the periphery of capsules also undergo apoptosis. Collectively, our results suggest that spread plasmatocytes release one or more factors that induce apoptosis of granular cells, and that this response is important in the final phases of capsule formation.     

Monoclonal antibodies bind distinct classes of hemocytes in the moth Pseudoplusia includens

Insect hemocytes have historically been identified on the basis of morphology, ultrastructure and hypothesized function. Among insects in the order Lepidoptera, five hemocyte classes are usually recognized: granular cells, plasmatocytes, spherule cells, oenocytoids and prohemocytes. We have generated a panel of monoclonal antibodies (mAbs) against hemocytes of the moth Pseudoplusia includens. In this study, hemocyte identification using 16 different mAbs was compared to identification methods using morphological characters. Three main categories of mAb binding activity were identified: (1) mAbs that specifically labeled only one morphological class of hemocytes, (2) mAbs that labeled granular cells and spherule cells, and (3) mAbs that labeled plasmatocytes and oenocytoids. With one exception, none of the antibodies bound to other tissues in P. includens. However, certain mAbs that specifically labeled granular cells and/or spherule cells in separated hemocyte populations also labeled plasmatocytes co-cultured with granular cells or cultured in granular cell conditioned medium. Overall, our results suggest that granular cells are antigenically related to spherule cells, and that plasmatocytes are antigenically related to oenocytoids. The use of mAbs as hemocyte markers are discussed.     

Insect hemocytes and their role in immunity

The innate immune system of insects is divided into humoral and cellular defense responses. Humoral defenses include antimicrobial peptides, the cascades that regulate coagulation and melanization of hemolymph, and the production of reactive intermediates of oxygen and nitrogen. Cellular defenses refer to hemocyte-mediated responses like phagocytosis and encapsulation. In this review, we discuss the cellular immune responses of insects with emphasis on studies in Lepidoptera and Diptera. Insect hemocytes originate from mesodermally derived stem cells that differentiate into specific lineages identified by morphology, function, and molecular markers. In Lepidoptera, most cellular defense responses involve granular cells and plasmatocytes, whereas in Drosophila they involve primarily plasmatocytes and lamellocytes. Insect hemocytes recognize a variety of foreign targets as well as alterations to self. Both humoral and cell surface receptors are involved in these recognition events. Once a target is recognized as foreign, hemocyte-mediated defense responses are regulated by signaling factors and effector molecules that control cell adhesion and cytotoxicity. Several lines of evidence indicate that humoral and cellular defense responses are well-coordinated with one another. Cross-talk between the immune and nervous system may also play a role in regulating inflammation-like responses in insects during infection.     

Parasitism by Cotesia plutellae alters the hemocyte population and immunological function of the diamondback moth, Plutella xylostella

Cotesia plutellae, a solitary endoparasitoid wasp, parasitizes the diamondback moth, Plutella xylostella, and induces host immunosuppression and lethality in the late larval stage. This study focused on changes of cellular immunity in the parasitized P. xylostella in terms of hemocyte composition and cellular functions. In third and fourth instar larvae of nonparasitized P. xylostella, granular cells represented the main hemocyte type (60-70%) and plasmatocytes were also present at around 15% among the total hemocytes. Following parasitization by C. plutellae, the relative proportions of these two major hemocytes changed very little, but the total hemocyte counts exhibited a significant reduction. Functionally, the granular cells played a significant role in phagocytosis based on a fluorescence assay using fluorecein isothiocyanate-labeled bacteria. The phagocytic activity of the granular cells occurred as early as 5 min after incubation with the bacteria, and increased during the first 40 min of incubation. The parasitism by C. plutellae significantly inhibited phagocytosis of the granular cells. Plasmatocytes also exhibited minor phagocytic activity. Moreover, plasmatocyte phagocytosis was not inhibited by parasitism. On the other hand, hemocyte-spreading behavior in response to pathogen infection was significant only for plasmatocytes, which exhibited a characteristic spindle shape upon infection. A significant spreading of the plasmatocytes was found as early as 5 min after pathogen incubation and their ratio increased during the first 40 min.An insect cytokine, plasmatocyte-spreading peptide 1 (PSP1) from Pseudoplusia includens, was highly active in inducing plasmatocyte-spreading behavior of P. xylostella in a dose-dependent manner. P. xylostella parasitized by C. plutella was significantly inhibited in plasmatocyte-spreading in response to an active dose of PSP1. An in vivo encapsulation assay showed that the parasitized P. xylostella could not effectively form the hemocyte capsules around injected agarose beads. This research demonstrates that the parasitism of C. plutellae adversely affects the total hemocyte populations in number and function, which would contribute to host immunosuppression.     

Integrin β1 subunit from Ostrinia furnacalis hemocytes: Molecular characterization, expression, and effects on the spreading of plasmatocytes

When lepidopteran larvae are infected by a large quantity of pathogens or parasitized by nonadaptive parasitoids, hemocytes in the hemocoel will encapsulate these foreign invaders. Cellular encapsulation requires hemocytes, particularly plasmatocytes, to change their states from nonadhesive, spherical cells into adhesive, spreading cells. However, it is unclear how the changes of plasmatocytes are regulated. Here we report that the integrin β1 subunit from hemocytes of Ostrinia furnacalis (Ofint β1) plays an important role in regulating the spreading of plasmatocytes. The full length cDNA sequence (4477 bp) of Ofint β1 was cloned from hemocytes. Phylogenetic analysis showed that Ofint β1 belonged to the integrin βPS family of Drosophila melanogaster with highest sequence identity (78.7%) to the β-integrin of Pseudoplusia includens. Structural analysis of the deduced amino acid sequence indicated that Ofint β1 had similar functional domains to known β-integrins in other lepidopteran insects. RT-PCR, Northern blotting, Western blotting and immunohistochemical analyses showed that OfINT β1 was expressed mainly in hemocytes, especially in plasmatocytes, and weakly in fat body, Malpighian tubes and epidermis. After hemocytes had spread onto slides, fewer antibodies to OfINT β1 bound to the surface of plasmatocytes. Furthermore, anti-OfINT β1 serum clearly inhibited the spreading of plasmatocytes. Together these results indicate that OfINT β1 may play an important role in regulating the spreading of plasmatocytes.     

Hematopoiesis in larval Pseudoplusia includens and Spodoptera frugiperda

Maintenance of circulating hemocytes in larval Lepidoptera has been attributed to both mitosis of hemocytes already in circulation and the release of hemocytes from hematopoietic organs. In this study, we compared hematopoiesis in the noctuids Pseudoplusia includens and Spodoptera frugiperda. For both species, hemocyte densities per microl of blood increased with instar. Differential hemocyte counts indicated that plasmatocytes were the most abundant hemocyte type during early instars but granular cells were the most abundant hemocyte type in the last instar. Hematopoietic organs were located in the meso- and metathorax of S. Frugiperda and P. Includens. These organs contained large numbers of hemocytes in S. Frugiperda, but contained few hemocytes in P. Includens. The majority of the hemocytes recovered from hematopoietic organs were identified as plasmatocytes. Using hemocyte type-specific markers and bromodeoxyuridine (BrdU) incorporation experiments, we determined that all hemocyte types with the exception of oenocytoids synthesize DNA. BrdU labeling indices for both species also fluctuated with the molting cycle. Ligation experiments suggested that hematopoietic organs are an important source of circulating plasmatocytes in S. Frugiperda but not in P. Includens. Injection of heat killed bacteria into larvae induced higher levels of BrdU labeling than injection of sterile saline, suggesting that infection and wounding induce different levels of hemocyte proliferation. Arch.     

Immune challenges trigger cellular and humoral responses in adults of Pterostichus melas italicus (Coleoptera,Carabidae)

The present study focuses on the ability of Pterostichus melas italicus Dejean to mount cellular and humoral immune responses against invading pathogens. Ultrastructural analyses revealed the presence of five morphologically distinct types of hemocytes: prohemocytes, plasmatocytes, granulocytes, oenocytoids and macrophage-like cells. Differential hemocyte counts showed that plasmatocytes and granulocytes were the most abundant circulating cell types and plasmatocytes exhibited phagocytic activity following the latex bead immune challenge. Macrophage-like cells were recruited after the immune challenge to remove exhausted phagocytizing cells, apoptotic cells and melanotic capsules formed to immobilize the latex beads. Total hemocyte counts showed a significant reduction of hemocytes after latex bead treatment. Phenoloxidase (PO) assays revealed an increase of total PO in hemolymph after immune system activation with lipopolysaccharide (LPS). Moreover, the LPS-stimulated hemocytes showed increased protein expression of inducible nitric oxide synthase, indicating that the cytotoxic action of nitric oxide was engaged in this antimicrobial collaborative response. These results provide a knowledge base for further studies on the sensitivity of the P. melas italicus immune system to the environmental perturbation in order to evaluate the effect of chemicals on non-target species in agroecosystems.     

Cross-protection experiments with parasitoids in the genus Microplitis (Hymenoptera: Braconidae) suggest a high level of specificity in their associated bracoviruses

The immunological and developmental effects of bracoviruses (BVs) from three parasitoids in the genus Microplitis (Braconidae: Microgastrinae) were compared in the hosts Pseudoplusia includens and Heliothis virescens (Lepidoptera: Noctuidae). Southern blotting experiments indicated that viral DNAs from Microplitis demolitor bracovirus (MdBV) cross-hybridized with viral DNAs from Microplitis croceipes bracovirus (McBV) and Microplitis mediator bracovirus (MmBV) under conditions of high stringency. Injection of calyx fluid plus venom from each parasitoid species dose-dependently delayed development of P. includens and H. virescens. Each virus also inhibited pupation of P. includens but not H. virescens. In situ hybridization experiments indicated that MdBV and McBV persistently infect hemocytes in both hosts while MmBV persistently infects hemocytes in P. includens but not H. virescens. While MdBV infection induced a loss of adhesion by most plasmatocytes, McBV and MmBV infection induced a loss of adhesion in less than 50% of cells. Cross-protection experiments indicated that calyx fluid plus venom from one species usually protected progeny of another species from encapsulation but did not always promote successful development.     

两种金小蜂毒液对菜粉蝶蛹血细胞延展、存活及包囊作用的影响

活体微注射测定结果表明,将0.5毒囊当量（venom reservoir equivalent, VRE）的蝶蛹金小蜂毒液注射于其寄主菜粉蝶蛹体内,注射后4～24 h寄主浆血细胞和颗粒血细胞的延展、存活和对Sephadex A50微珠的包囊能力显著下降;以0.002～0.02 VRE/μL的该蜂毒液处理其离体寄主血细胞均能产生同样的生理效应。该毒液抑制90%菜粉蝶蛹浆血细胞和颗粒血细胞延展的浓度各为0.00076 VRE/μL和0.00804 VRE/μL。可见,蝶蛹金小蜂毒液能显著抑制其寄主血细胞的延展和包囊作用,并导致血细胞的死亡。然而,同样条件下丽蝇蛹集金小蜂毒液对其非自然寄主菜粉蝶蛹的血细胞延展、存活和包囊作用则无任何效应。可见,寄生蜂毒液的生理作用具有明显的寄主特异性。     

A NOVEL TYPE OF HEMOCYTES LOCALIZING MELANIZATION WITH HIGH‐SPREADING BEHAVIOR IN MYTHIMNA SEPARATA

Lepidopteran larvae show a cellular response to invading foreign substances that are larger than hemocytes, for example, parasitoid eggs or larvae. This response is called hemocyte encapsulation and is often accompanied by phenoloxidase (PO)‐catalyzed melanization. In the present study, we artificially transplanted endoparasitoid larvae and small glass fragments into the hemocoel of the common armyworm, Mythimna separata. We observed that the host larva showed a cellular response and that, 2–4 h after transplantation, melanin formation was spatially confined to the surface of the encapsulated substances. We further noted that specific morphological hemocytes surrounded by melanin formation became attached to the surface of the foreign substances. We designated these hemocytes hyperspread cells (HSCs) on the basis of their specific characteristics and circumferential spread. We confirmed the occurrence of prophenoloxidase (PPO)/phenoloxidase (PO) on the periphery of the HSCs and in the substance secreted around the HSCs by using anti‐PPO antibody. We were unable to detect PPO‐mRNA in HSCs by using in situ hybridization, although we showed that oenocytoids contained PPO‐mRNA and PPO protein. We used light microscopy and scanning electron microscopy to discriminate five main types of circulating M. separata hemocytes. We observed that HSCs differed from plasmatocytes, but spread out well. Further, during the encapsulation process, HSCs appeared to provide a localized melanization spot on the surface of foreign invaders.     

Influence of calcium on Manduca sexta plasmatocyte spreading and network formation

Plasmatocytes are a class of insect hemocytes important in the cellular defense response. In some species, they are phagocytic, protecting the insect from smaller pathogens. In many insects, they work in concert with other hemocytes (particularly other plasmatocytes and granular cells) to form nodules and to encapsulate foreign material. To perform these functions, plasmatocytes attach to, spread on, and surround suitable targets. Because of their importance, because we had previously observed that prolonged incubation of hemocytes in solutions containing the divalent cation chelator ethylenediaminetetraacetic acid (EDTA) inhibited plasmatocyte spreading, and because of the importance of divalent cations in many immune-related functions, we investigated the effect of calcium and magnesium on spreading of plasmatocytes from fifth instar Manduca sexta larvae. On glass slides, plasmatocytes spread more quickly and elongated in Grace's medium containing 5 mM calcium, compared to calcium-free medium. In the presence of calcium, plasmatocyte adhesion, spreading, and network formation were not visibly different in magnesium-free and magnesium-containing Grace's medium. Using immunomicroscopy with a monoclonal antibody specific for plasmatocytes, we measured the length and width of plasmatocytes incubated with several different concentrations of calcium. Plasmatocyte length positively correlated with calcium concentration to 5 mM (maximum concentration tested and approximately the hemolymph concentration). Mean plasmatocyte width was less in 0 and 5 mM calcium than in 0.05 or 0.5 mM calcium. On plastic, hemocytes survived longer than on glass (they survived beyond 24 h) and, in 5 mM calcium, formed an extensive network readily visible by phase-contrast microscopy. This network was never as extensive in the absence of calcium. Network formation in the absence of magnesium, but presence of calcium, resembled network formation in standard Grace's medium.     

The ectoparasitic wasp Nasonia vitripennis (Walker) (Hymenoptera: Pteromalidae) differentially affects cells mediating the immune response of its flesh fly host,Sarcophaga bullata Parker (Diptera: Sarcophagidae)

In this study, we examined cellular immune responses in the flesh fly, Sarcophaga bullata, when parasitized by the ectoparasitoid Nasonia vitripennis. In unparasitized, young pharate adults and third instar, wandering larvae of S. bullata, four main hemocyte types were identified by light microscopy: plasmatocytes, granular cells, oenocytoids, and pro-hemocytes. Parasitism of young pharate adults had a differential effect on host hemocytes; oenocytoids and pro-hemocytes appeared to be unaltered by parasitism, whereas adhesion and spreading behavior were completely inhibited in plasmatocytes and granular cells by 60 min after oviposition. The suppression of spreading behavior in granular cells lasted the duration of parasitism. Plasmatocytes were found to decline significantly during the first hour after parasitism and this drop was attributed to cell death. Melanization and clotting of host hemolymph did not occur in parasitized flies, or the onset of both events was retarded by several hours in comparison to unparasitized pharate adults. Hemocytes from envenomated flies were altered in nearly identical fashion to that observed for natural parasitism; the total number of circulating hemocytes declined sharply by 60 min post-envenomation, the number of plasmatocytes declined but not granular cells, and the ability of plasmatocytes and granular cells to spread when cultured in vitro was abolished within 1 h. As with parasitized hosts, the decrease in plasmatocytes was due to cell death, and inhibition of spreading lasted until the host died. Isolated crude venom also blocked adhesion and spreading of these hemocyte types in vitro. Thus, it appears that maternally derived venom disrupts host immune responses almost immediately following oviposition and the inhibition is permanent. The possibility that this ectoparasite disables host defenses to afford protection to feeding larvae and adult females is discussed.     

亚洲玉米螟幼虫血细胞的包囊行为

根据光镜和电镜观察结果,将亚洲玉米螟(Ostrinia furnacalis)幼虫血细胞分为粒细胞、浆细胞、类绛色细胞、原血细胞和球形血细胞五类。调查了幼虫的血细胞总数(THC)和各类血细胞数量(DHC)的变化情况。从三龄末期到五龄第五天期间,幼虫的THC在蜕皮前后会下降,蜕皮后约12h降到最低点,然后又慢慢回升。在五龄幼虫前5d期间,浆细胞在前第三天呈增加趋势,之后开始下降,而粒细胞呈相反趋势。浆细胞和粒细胞具有附着延展性,它们可以附着在载玻片表面,但延展能力不同。血细胞可以迅速黏附在外源物如葡聚糖凝胶珠表面形成包囊,部分包囊会发生黑化现象。体外培养条件下,血细胞也可以形成包囊,其结构与体内形成的包囊差异不大。     

Correlation of light and electron microscopic hemocyte structue in the dictyoptera

As part of program of research into insect cellular immunity, an integrated light and electron microscopic study of the hemocytes of seven members of the Order Dictyoptera has been made. In fresh hemolymph, five cell types, the prohemocytes, plasmatocytes, granular cells, spherule cells and cystocytes, are easilv distinguished. However, in thick Araldite sections and in thin sections in the electron microscope it is sometimes difficult to identify the various cell types. The reasons for this difficulty are discussed. Granules with a microtubular substrcture are found in the plasmatocytes, spherule cells and cystocytes. In the plasmatocytes these granules have a different ultrastructure than those in the spherule cells and cystocytes. The in vitro fragility of these granules in both the spherule cells and cystocytes during coagulation partially explains the previous confusion in distinguishing these two cell types. Evidence is presented which indicates that the plasmatocytes, granular cells and spherule cells represent a developmental series originating from the prohemocytes. Where exactly the cystocytes are derived from is unknown.     

颈双缘姬蜂毒液对寄主小菜蛾的免疫抑制作用

对颈双缘姬蜂Diadromus collaris (Gravenhorst)及其毒液引起寄主小菜蛾Plutella xylostella的一些生理效应进行了研究。结果表明,颈双缘姬蜂寄生寄主后可引起寄主小菜蛾蛹总血细胞及浆血细胞和颗粒血细胞数量的上升。寄生后1天观察,血细胞延展行为受到影响,表现在颗粒血细胞放射状丝的产生及浆血细胞伪足的形成受到抑制。通过毒液对寄主离体幼虫血细胞延展行为、形态及活性影响的研究,发现毒液抑制了寄主离体浆血细胞的延展,但对颗粒血细胞的影响不明显;毒液引起寄主浆血细胞和颗粒血细胞的破裂和死亡,毒液对寄主幼虫血淋巴酚氧化酶活性有一定的抑制作用,当反应至40、60及80 min时,毒液处理和未经毒液处理的寄主血淋巴在490 nm处的吸光值差异比较明显。对毒液蛋白成分的聚丙烯酰胺凝胶电泳分析发现,毒液中有9种多肽,分子量介于9～50.2 kD,其中50.2、30.5、28.2、25.1 和12.6 kD的多肽含量较高, 与其他蜂毒液的一些作用已知的蛋白条带相似,因而推测它们同样具有免疫及发育抑制作用。结果证明颈双缘姬蜂毒液能破坏寄主细胞及体液因子调节的免疫反应。