Insect Hemocytes Research Articles

Affiliation of large numbers of fibroblasts with larval muscle fibers

Muscle fibers from fourth and fifth instar caterpillars were examined with scanning and thin section electron microscopy. Scanning micrographs showed that early fifth instar specimens had a population of cells lying beneath the basal lamina over the surface of the muscle fiber and in conjunction with tracheoles and nerves. At least two cell types were present. One type could be categorized as tracheoblasts by their close association with the tracheoles and the presence of taenidia within the tracheoblast cytoplasm in sectioned material. A second cell type, characterized by long filamentous processes, contained extensive rough endoplasmic reticulum and cisternae swollen with an electron-dense substance similar in appearance to the basal lamina. This ultrastructural appearance is characteristic of vertebrate fibroblasts and certain types of insect hemocytes. Early and late fourth instar specimens had few cells on their muscle fiber surfaces. Measurements of the basal lamina thickness were taken from thin sections of non-digested muscle fibers of early fourth, late fourth, and early fifth instar animals. The results showed that the basal lamina underwent a large increase in thickness between the fourth and fifth instars. The proliferation of cells which appeared to be in an actively synthesizing state paralleled the increase in basal lamina thickness. This suggests the hypothesis that these cells are active in connective tissue formation, and contribute to the formation of the basal lamina that lies over both them and the muscle fiber.

Effect of substratum wettability and charge on adhesion in vitro and encapsulation in vivo by insect haemocytes

The circulating leucocytes of insects, the haemocytes, adhere to and encapsulate foreign material that enters the insect's body cavity. The thickness of the capsule depends not only on the insect species but also on the nature of the object concerned, a fact that is of great importance to invading parasites and pathogens. In this paper, some of the factors that may stimulate haemocyte adhesion and subsequent encapsulation of the object have been investigated using abiotic materials with surfaces of different charge and wettability. The negativity and wettability of surfaces of polystyrene beads and plates can be increased by pretreatment with acid, and adhesion of haemocytes to these modified surfaces has been examined in vivo and in vitro. A similar proportion of haemocytes of the locust Schistocerca gregaria adhere to the plates in vitro, irrespective of the changes in charge and wettability, but the adhesion of haemocytes of the cockroach Periplaneta americana is proportional to the increases in both parameters. These differences in cell behaviour are reflected in vivo: cockroach haemocytes form thicker capsules around more hydrophilic and more negatively charged polystyrene beads, while locust cells encapsulate both types of surface to the same, minimal, degree. Positively and negatively charged Sepharose beads are encapsulated more thickly than are neutral beads in cockroaches; negatively charged Sepharose beads are not encapsulated at all in locusts. There are thus obvious differences between the two species in the ways in which their haemocytes respond to these modified abiotic surfaces. It is suggested that capsule thickness in vivo depends on the initial cell-substratum contact; different surfaces stimulate the cell to different extents, which in turn causes variations in the recruitment of other cells to the capsule.

Drosophila larval fat body surfaces : Changes in transplant compatibility during development.

The hemocytes oftu-Sz ts melanotic tumor larvae ofDrosophila melanogaster encapsulate heterospecific and surface-modified homospecific tissue implants, but do not encapsulate unmodified homospecific implants (R. Rizki and Rizki 1980). In the present study we usedtu-Sz ts hosts to assay changes in larval fat body surfaces during development. Donor fat bodies from various ages of larvae were accepted (remained unencapsulated) intu-Sz ts hosts whereas fat bodies from donors with everted spiracles and all subsequent stages of development that were tested were rejected (encapsulated). Since the demarcation between acceptance and rejection by thetu-Sz ts blood cells did not coincide with the gross morphological changes that appear in the fat body during metamorphosis (dissolution of the basement membrane and dispersal of the freed fat body cells at pupation), we compared acceptable and nonacceptable fat body surfaces by three other methods. Fat body surface ultrastructure was examined, fat bodies were treated with fluorescein isothiocyanate-conjugated lectins, and fat body surfaces were reacted with a monoclonal antibody specific for basement membrane. These approaches did not uncover fat body surface changes associated with eversion of the anterior spiracles, suggesting that recognition of tissue surface heterogeneities by the insect hemocytes exceeds the resolving power of the other three methods. However, the monoclonal antibody fails to bind to the basement membrane ofD. virilis larvae, whose fat body is always rejected intu-Sz ts hosts. This supports our suggestion that the molecular architecture of the basement membrane may be important in eliciting the encapsulation response.

Octopamine-sensitive adenylate cyclase in haemocytes of the forest tent caterpillar, Malacosoma disstria Hubner (Lepidoptera: Lasiocampidae)

Injection of octopamine into the haemocoel of third instar larvae and pupae of the forest tent caterpillar, Malacosoma disstria, results in a rapid elevation of haemolymph cyclic AMP levels. The elevated cyclic AMP levels are due, at least in part, to the action of octopamine on haemocytes, as evidenced by the octopamine-induced stimulation of adenylate cyclase in an established haemocyte cell line (Md 66). The effect is simulated by the adrenergic agonist, synephrine, and inhibited by the α-adrenergic blocking agent, phentolamine. The insect haemocyte receptor–enzyme system resembles equivalent vertebrate systems in being activated by sodium fluoride and requiring GTP for expression of effector-induced activation of the enzyme.

Insect hemocytes: Cells adapted to anaerobiosis

Hemocytes from a free-living insect could be grown under strictly anoxic conditions. Their anaerobic metabolism was studied in vitro in such a cellular system.

Insect Hemocytes: Development, Forms, Functions, and Techniques.A. P. Gupta

Insect Hemocytes: Development, Forms, Functions, and Techniques.A. P. Gupta

In vivo insect hemocyte destruction by UV-irradiated Chilo iridescent virus

Large inoculum of infective or UV-irradiated Chilo iridescent virus (CIV) caused an early drastic cytopathy of the larval hemocytes of the greater wax moth, Galleria mellonella. Cytopathy of the hemocyte could be detected 2 hr after virus injection at 25°C and most hemocytes were completely destroyed within 8 hr after virus injection. Heat-treated CIV did not produce any cytopathy of Galleria hemocytes. Antiserum to CIV neutralized this cytopathic effect, while that to Tipula iridescent virus did not demonstrate any inhibitory effect. Actinomycin D, mitomycin C, and puromycin did not inhibit hemocyte destruction by UV-irradiated CIV. Galleria larva injected with a large dose of UV-irradiated CIV progressively became stunted with a decrease of body weight and did not regain its hemocyte numbers. Hemocytes infected with CIV were not destroyed by any additive injection of a large dose of UV-irradiated CIV.

Insect haemocyte preparation techniques for sem

Insect haemocyte preparation techniques for sem

The cystacanth of Moniliformis moniliformis (Bremser, 1811) and its relationship with the haemocytes of the intermediate host (Periplaneta americana).

The nature of the capsule of the cystacanth of Moniliformis moniliformis was studied. Fat body, tracheal system, Malpighian tubules and haemocytes of the host are intimately associated with the cystacanth. The capsule of the cystacanth is cellular. The cells encapsulating the middle and late acanthellae are granular haemocytes of the host, whereas those encapsulating the cystacanth are agranular. Degranulation of haemocytes after encapsulation of the cystacanth recalls the common method for encapsulation of parasites of insect haemocytes, but a difference between the two kinds of encapsulation is that the capsule around the cystacanth does not bring about melanization nor destroys the parasite. It is suggested that the surface of the parasite may have inhibitors against enzyme systems causing melanization. Host haemocytes are also found in the capsular fluid. The infiltration of haemocytes into the capsule may be in response to the release of metabolites by the parasites. It is also suggested that the haemocytes may detoxify the parasitic metabolities. Rarely, capsules containing two parasites were observed. In one of the cases, one of the two cystacanths already possessed its own capsule. This finding, together with the cellular nature of the capsule, suggests that the capsule is of host origin.

Differential adhesion of the haemocytes of Leucophaea maderae (Blattaria) to a glass surface

A system for the study of insect haemocytes in vitro is described. The system was used to analyse the adhesive properties of the haemocytes of the cockroach, Leucophaea maderae. The two main types of haemocytes, plasmatocytes and granulocytes, showed considerable differences in adhesive properties, which allowed the production of nearly homogeneous monolayers consisting of either plasmatocytes or granulocytes. The much stronger adhesion of the plasmatocytes is discussed in relation to their role in phagocytosis and encapsulation.

Cellular defense reactions of insect hemocytes in vivo: Nodule formation and development in Galleria mellonella and Pieris brassicae larvae

Galleria mellonella and Pieris brassicae larvae were injected with a standardized dose of killed Bacillus cereus and other bacteria and the reactions of hemocytes followed in the first 24 hr by dissection and histology. Nodules formed in all insects injected with nonpathogens, but a pathogen, Staphylococcus aureus, failed to provoke this reaction. Within 5 min, clumps consisting of granular hemocytes, plasmatocytes, and bacteria were found attached to the internal surfaces of the insects. In the following hours, the cells comprising the clumps broke down and merged with a melanizing acellular substance, and the necrosing masses became encapsulated by plasmatocytes to form mature nodules. The role of granular hemocytes in the formation of the initial cell/bacteria aggregates is discussed along with the possible importance of nodules to the cellular defense reactions of insects.

Cellular defense reactions of insect hemocytes in vitro: Phagocytosis in a new suspension culture system

In a newly developed short-term culture system the plasmatocytes of Galleria mellonella, Pieris brassicae, Calliphora erythrocephala, and Periplaneta americana are the most active cell types in the phagocytosis of latex, chick erythrocytes, and certain bacteria. The granular cells of G. mellonella and the spherule cells of P. brassicae also phagocytose these test particles to a limited extent. This culture system is described, together with the appearance of ingested particles in preparations of living cells, and in fixed and stained monolayers. In culture, the hemocytes clump together due to cell instability in vitro and to the presence of uningested particles. This clumping reaction may be similar to nodule formation observed in vivo in these insects.

Effects of trematode infection on the hemolymph of aquatic insects

The effects of trematode infection on the hemolymph proteins of mayfly naiads ( Hexagenia recurvata) and alderfly larvae ( Sialis sp.) were studied using polyacrylamide gel disc electrophoresis. One-to 14-day experimental infections with Allopodocotyle lepomis cercariae were associated with depletion of total hemolymph protein in both species. Protein was lost from all fractions of Hexagenia hemolymph except the fastest migrating one. Two fractions of Sialis hemolymph which showed significant reduction in protein contained tyrosinase activity which also decreased during infection. Tyrosinase was also located in one-fifth to one-half of the hemocytes of both insects. These results suggest that the tyrosinase involved in the melanization of trematode larvae within Hexagenia is confined to the hemocytes, whereas in Sialis the hemolymph tyrosinase may also play an important role.

In vitro phagocytosis of bacteria by insect blood cells.

PHAGOCYTOSIS is an important component of vertebrate cellular immunity and involves several stages which have been described in detail previously1,2. In contrast, little is known about this process in insects even though recent work3,4, on intracellular killing of bacteria indicates that phagocytosis probably plays an important role in the defence reactions of insects. In vertebrates, the development of in vitro techniques has greatly facilitated studies on phagocytosis, but in insects such techniques are not generally available for studying the blood cells (haemocytes) in strictly controlled conditions. Here we describe for the first time the stages in the phagocytosis of bacteria by insect haemocytes, using a newly developed culture technique.

Free amino acids in the haemocytes and plasma of the larva of Calliphora vicina.

ABSTRACT Free amino acid concentrations have been measured in haemolymph samples taken from 3rd instar larvae of the blowfly, Calliphora vicina, at various stages prior to pupariation. The amino acids found in the haemocyte fraction only accounted for 6% of the total free amino acid concentration of the haemolymph. However, a high percentage of the dicarboxylic amino acids, glutamate and aspartate, 62% and 69% respectively, appeared to be sequestered in the haemocyte fraction at 72 h prior to pupariation. The percentage of the other individual amino acids found in the haemocyte fraction represented less than 10% of their amount in whole haemolymph. It is proposed that these results, together with the increase in the haemocyte levels of glutamate observed after injecting larvae with saline containing glutamate, are one of the first indications of a homeostatic function of insect haemocytes with respect to haemolypmh amino acids.

The hemocytes of an lsopod Ligia exotica Roux.

AbstractThe morphological features of the hemocytes of the crustacean Ligia exotica are similar to hemocytes of insects and millipedes. Jones system of hemocyte classification is extended to crustacean hemocytes. As in insects, seven classes of hemocytes, identified as prohemocytes, plasmatocytes, granular hemocytes, cystocytes, oenocytoids, spherule cells and adipohemocytes, occur. The prohemocytes can be subdivided into five categories that probably represent the precursor of major cell types. The structural and chemical features of other major cell classes are distinct and support the concept of Jones ('62) that these types might have different lineages and might not be capable of transforming into one another. Some of the prohemocytes, plasmatocytes and granular hemocytes are amoeboid. Cystocytes do not bring about any visible plasma coagulation similar to their counterpart in millipedes. Oneocytoids and adipohemocytes are rare. Plasmatocytes, cystocytes and oenocytoids occur in conglomerates, the significance of which is discussed. The cell types are compared with those of the hemocytes of other crustaceans. It is suggested that the nomenclature based on morphological characters is more suited for crustacean hemocytes than a nomenclature based on behavioural and physiological characters.

The hemocytes of a scorpion Palamnaeus swammerdami.

AbstractSome of the cytological characteristics of the hemocytes of the scorpion, Palamnaeus swammerdami, were studied. The morphological features of the arachnid hemocytes were observed to be similar to those of hemocytes of insects, millipedes and isopods. Jones' system of hemocyte classification was extended to the arachnid hemocytes. The six classes of hemocytes indentified in the scorpion correspond to prohemocytes, plasmatocytes, granular hemoocytes, cystocytes, spherule cells and adipohemocytes of insects. A cell type comparable to oenocytoids of insects and crustaceans is absent. The prohemocytes can be subdivided into four categories that probably represent the precursors of the major cell types. The cytological characteristics of the major cell classes and the occurrence of the miniatures of some of these major cell types support the concept of Jones (62) that these cell types might have different cell lineages and might not be capable of transforming into one another. Some of the prohemocytes, plasmatocytes and granular hemocytes were amoeboid. The nature of the granules and the vacuoles of plasmatocytes and granular hemocytes were compared with the granules and vacuoles of corresponding hemocytes of other arthropods. Cystocytes did not bring about any visible coagulation similar to their counterparts in millipedes and crustaceans. Plasmatocytes, granular hemocytes and spherule cells were observed to occur in conglomerates. The cell types noted in the present study were compared with the hemocytes of other arachnids.

A reappraisal of insect haemocyte classification by the examination of blood from fifteen insect orders.

A simplified insect haemocyte classification has been formulated by a light microscopic examination of the haemolymph of insects from fifteen Orders. Six cell types or developmental stages can be distinguished: (1) Prohaemocytes, (2) Plasmatocytes, (3) Granular Cells, (4) Spherule Cells, (5) Cystocytes, and (6) Oenocytoids. The structure and occurrence of these haemocytes are described together with the structural variations which occur in each cell type. Due to considerable overlap in structure and the presence of numerous intermediates the six cell types may represent different developmental and/or functional stages of one basic cell type. The available evidence for this unitarian hypothesis is discussed.

The hemocytes of a millipede, Thyropygus poseidon.

Some of the cytological and histochemical characteristics of hemocytes of Thyropygus poseidon were studied. Jone's system of classification of hemocytes was extended to millipede hemocytes. Seven classes were identified as prohemocytes, plasmatocytes, granular hemocytes, cytocytes, oenocytoids, spherule cells and adipohemocytes. Oenocytoids were rarely found. All cells except prohemocytes and oenocytoids sequestered oxidized products of catechol. Results of the histochemical tests for proteins also suggest that these cells sequester plasma proteins. Cytocytes do not bring about any visible coagulation similar to the hemocytes of some hemipteran insects. Granular hemocytes contain phenol-oxidase capable of oxidising tyrosine as well as catechol, similar to the hemocytes of crustaceans. In this respect millipede hemocytes differ from insect hemocytes. It is suggested that these cell types may represent distinct cell lines and may not represent transformation of one cell type to another.

Comparative biochemistry of phagocytizing insect hemocytes

1. 1. The respiratory increment associated with phagocytosis by mammalian leukocytes is absent in Blaberus craniifer hemocytes. 2. 2. The glycolytic pathway provides energy for particle engulfment in both mammalian leukocytes and B. craniifer phagocytes. 3. 3. Hexose monophosphate pathway activity of roach hemocytes, unlike mammalian cells, is not stimulated by phagocytosis. 4. 4. Nitroblue tetrazolium reduction, characteristic of mammalian phagocytes, is absent in B. craniifer cells. 5. 5. B. craniifer hemocytes utilize antimicrobial systems other than the myeloperoxidase-H 2O 2-halide system of mammals.