Comparative study of hemocytes and associated cells of some medically important dipterans

The aim of this work is to study, characterize, and compare different morphological types of hemocytes of Glossina austeni, G. morsitans, Calliphora erythrocephala, Stomoxys calcitrans, Lucilia sericata, Aedes aegypti, and Culex quinquefasciatus. This information is intended to provide a basis for future studies of the cellular defense mechanisms of these dipterans. Seven morphological types of hemocytes were identified by phase-contrast optics: prohemocytes, plasmatocytes, thrombocytoids, granulocytes, adipohemocytes, oenocytoids, and spindle cells of various sizes. Adipohemocytes are difficult to distinguish from both fat body cells and granulocytes. All seven cell types are not present in every species. For example, thrombocytoids and spindle cells were not found in A. aegypti or C. quinquefasciatus, and oenocytoids were observed only in A. aegypti, C. quinquefasciatus, and in C. erythrocephala. In addition to the hemocytes, fat body cells and nephrocytes are also freely present in the hemolymph of some species but may have gained access to the blood during the bleeding process. With electron microscopy and with thick plastic sections of G. austeni hemolymph, only nephrocytes, plasmatocytes, granulocytes, and spindle cells were identified with any certainty, and the large spindle cells are morphologically very different from those found in the other dipterans. They are rigid cells supported by microtubules running throughout their entire length. These hemocytes are present in large numbers only in newly emerged flies, they are absent in larvae and young pupae, and are rare in old adults. Their disappearance from the hemolymph of newly emerged Glossina appears to be a result of phagocytosis by the plasmatocytes.     

Nephrocytes of harvestmen, Leiobunum limbatum and L. rotundum

Nephrocytes are said to be able to take up substances from the hemolymph. In Opiliones, which were examined electron microscopically three different types of nephrocytes were found. Numerous large nephrocytes lie clustered between the muscles in the anterior region of the body. Smaller nephrocytes occur scattered throughout the opilionid body, often affixed to tracheoles. The third group, pericardial cells, are always attached to the heart wall by connective ligaments. All nephrocytes are surrounded by a thick basement membrane and their plasma membrane forms pedicels. Junctional complexes link the adjacent pedicels to form diaphragm-like slit-membranes, which form the entrance to an extracellular compartment. The cytoplasm of the nephrocytes contains many pinocytotic vesicles and tubular elements. Different types of large electron-dense and electron-lucent vesicles can be distinguished. Occasionally a large bundle of unmyelinated nerve fibers is enclosed by a pericardial cell. Morphological differences between the types of nephrocytes are described and the ultrastructural characteristics of the nephrocytes of harvestmen are compared with those of other arthropods. Functional aspects are discussed with respect to their ultrafiltration structures.     

The fine structural organization of sternal glands of pseudergates and workers in termites (Isoptera): a comparative survey

Thirty-nine species belonging to different families of termites are studied to give a comprehensive view of the evolution of the sternal glands. Several modifications occurring at cuticular and cytological levels are described in neuter castes. The outer epicuticle is always pierced by epicuticular pores. In advanced termites the epicuticular filaments greatly increase in number and length creating a thick layer. The pore canals gradually enlarge while the cuticle changes into a lattice structure lining an extracellular space in which the secretion is stored. Two classes of cells are present in basal termites (Mastotermitidae, Hodotermitidae, Termopsidae and Kalotermitidae) but their glandular structures greatly differ between families. A more complex organization with three classes of cells is found in the Serritermitidae and Rhinotermitidae. A regressive evolution occurs in the Termitidae where only two classes of cells are present. A dual nervous control (campaniform sensilla and neurosecretory fibers) is found in lower termites, except for the Hodotermitidae which have mechanosensory bristles. In the other families, neurosecretory fibers are lacking. A comparison with phylogenetic data is given. A more versatile role of sternal glands in neuter castes is hypothesized.     

培菌白蚁与鸡枞菌研究进展

培菌白蚁起源于非洲,蚁巢内具有复杂的社会分工.培菌白蚁依靠独特的蚁巢结构维持内部稳态和气体循环.菌圃是白蚁培育鸡枞菌的场所.鸡枞菌隶属于担子菌亚门,但其传播方式和生活史具有区别于其它担子菌的特点.鸡枞菌协助白蚁进行植物纤维的消化,白蚁则为鸡枞菌提供合适的生长环境,并控制鸡枞菌的遗传结构.培菌白蚁和鸡枞菌形成紧密的共生关...     

Colony-dependent sex differences in protozoan communities of the lower termite <Emphasis Type="Italic">Reticulitermes speratus</Emphasis> (Isoptera: Rhinotermitidae)

In many animals, sex differences in hormones, behavior, and immunity lead to differences in their gut microbial communities. One of the best-known examples of mutualistic symbiosis is that between lower termites and their intestinal protozoa. Although differences in the protozoan communities of different castes have been studied in lower termites, nothing is known about the sex differences in protozoan communities in neuter castes. Here, we show that termite workers have different protozoan communities according to sex depending on the colony. We investigated the communities of symbiotic protozoa living in lower termites, Reticulitermes speratus, and how they are affected by sex and caste. Workers had the largest numbers of protozoa, followed by soldiers, whereas reproductives (primary kings and secondary queens) had no protozoa. Workers showed colony-dependent sex differences in the total abundance of protozoa, whereas soldiers showed no such sex differences. There were significant sex effect and/or interaction effect between colony and sex in abundances of five species of protozoa in workers. Workers also showed significant sex differences and/or colony-dependent sex differences in proportion of six species of protozoa. These may result in sex differences in the host–symbiont interaction due to physiological or behavioral sex differences in workers that have not been recognized previously. This study has an important implication: although workers are not engaged in reproduction, their potential sex difference may affect various aspects of social interactions.     

Caste- and development-associated gene expression in a lower termite

Background  

Social insects such as termites express dramatic polyphenism (the occurrence of multiple forms in a species on the basis of differential gene expression) both in association with caste differentiation and between castes after differentiation. We have used cDNA macroarrays to compare gene expression between polyphenic castes and intermediary developmental stages of the termite Reticulitermes flavipes.     

Effects of juvenile hormone III on Reticulitermes flavipes: changes in hemolymph protein composition and gene expression

Termites express polyphenism during caste differentiation that is mostly undefined at the molecular level. Using the eastern subterranean termite, Reticulitermes flavipes Kollar, we wanted (1) to test juvenile hormone (JH) model assays for their ability to induce detectable molecular changes in worker termites and (2) to investigate hemolymph proteins and their corresponding genes during JH-induced soldier caste differentiation. Our results illustrate pronounced changes in two hemolymph proteins after JH treatment, as well as differences among several caste phenotypes. Significant increases in the expression of four genes encoding hemolymph proteins, including two vitellogenins and two hexamerins, were observed after JH exposure. These findings are the first to demonstrate such protein and gene expression changes during termite caste differentiation. These results also validate the utility of JH model assays for inducing detectable molecular changes in worker termites that have begun presoldier differentiation.     

The cytoskeleton of the ventral nephrocytes of Ceratitis capitata larva

Nephrocytes are cells involved in the regulation of the composition of the hemolymph and are characterized by peripheral finger-like projections delimiting a labyrinthine channel system. We have combined structural and immunological methods to examine the spatial distribution of microtubules and microfilaments in disseminated nephrocytes of Ceratitis capitata larva; the nephrocytes are scattered among the fat-body cells, close to the salivary glands. Actin filaments are localized in two discrete peripheral domains and microtubules are mostly concentrated in the cortical region. We discuss the possibility that these cytoskeletal elements, localized in the finger-like processes of the plasma membrane, are involved in maintaining the spatial architecture of the cell periphery and in modifying the junctional complexes that represent the entrance to the labyrinthine channel system.     

Soldier-specific modification of the mandibular motor neurons in termites

Social insects exhibit a variety of caste-specific behavioral tendencies that constitute the basis of division of labor within the colony. In termites, the soldier caste display distinctive defense behaviors, such as aggressively attacking enemies with well-developed mandibles, while the other castes retreat into the colony without exhibiting any aggressive response. It is thus likely that some form of soldier-specific neuronal modification exists in termites. In this study, the authors compared the brain (cerebral ganglion) and the suboesophageal ganglion (SOG) of soldiers and pseudergates (workers) in the damp-wood termite, Hodotermopsis sjostedti. The size of the SOG was significantly larger in soldiers than in pseudergates, but no difference in brain size was apparent between castes. Furthermore, mandibular nerves were thicker in soldiers than in pseudergates. Retrograde staining revealed that the somata sizes of the mandibular motor neurons (MdMNs) in soldiers were more than twice as large as those of pseudergates. The enlargement of MdMNs was also observed in individuals treated with a juvenile hormone analogue (JHA), indicating that MdMNs become enlarged in response to juvenile hormone (JH) action during soldier differentiation. This enlargement is likely to have two functions: a behavioral function in which soldier termites will be able to defend more effectively through relatively faster and stronger mandibular movements, and a developmental function that associates with the development of soldier-specific mandibular muscle morphogenesis in termite head. The soldier-specific enlargement of mandibular motor neurons was observed in all examined species in five termite families that have different mechanisms of defense, suggesting that such neuronal modification was already present in the common ancestor of termites and is significant for soldier function.     

Immunocytochemical localization of lysozyme in the nephrocytes of the harvestman, Leiobunum rotundum

Nephrocytes are cells involved in the metabolism of hemolymph components and are characterized by peripheral finger-like projections bordering a labyrinthine channel system. The antibacterial protein lysozyme was localized in anterior and pericardial nephrocytes of the harvestman, Leiobunum rotundum, by biochemical and immuno-gold postembedding-labelling techniques. Lysozyme-activity was demonstrated in experimental animals, challenged by Micrococcus luteus Grampositive bacteria. With SDS-electrophoresis, lysozyme was proved to be present in nephrocytes adjoining the heart, before studies at the electron microscopical level were carried out. The opilionid lysozyme was found to have a molecular weight of 14000 Da, thus belonging to the c-type lysozyme which is represented by hen-egg-white lysozyme. In untreated animals the presence of lysozyme could not be proved by electrophoresis. With immunogold labelling, however, lysozyme could be clearly localized in the vesicles of nephrocytes of untreated animals. Additional control experiments were carried out which revealed that the nephrocytes of L. rotundum in vivo are able to take up native lysozyme from an exterior medium. Therefore, the problem needs to be solved if nephrocyte vesicles transport lysozyme into or out of the cell.     

Pathological changes in the nephrocytes of the shore crab, Carcinus maenas, following injection of bacteria

The gills of Carcinus maenas were examined by light and electron microscopy following injection of either sterile saline or the bacteria Bacillus cereus and Moraxella sp., to determine any role(s) for the nephrocytes in the host defense reactions. The results showed that although intact bacteria were not sequestered to the nephrocytes, these cells were active in the removal of large quantities of cell debris from the hemolymph. Much of this material was derived from the breakdown of the hemocytes in response to the presence of bacteria and it's accumulation in the central vacuoles of the nephrocytes resulted in the degradation of these cells. It is proposed that while nephrocytes do not phagocytose intact bacteria, they augment the host defenses by clearing much of the hemocyte and associated bacterial debris from the gills, thus preventing blockage of the lamellar sinuses and subsequent impairment of respiration.     

Selection on defensive traits in a sterile caste – caste evolution: a mechanism to overcome life‐history trade‐offs?

SUMMARY During development and evolution individuals generally face a trade-off between the development of weapons and gonads. In termites, characterized by reproductive division of labor, a caste evolved—the soldiers—which is completely sterile and which might be released from developmental trade-offs between weapons and testes. These soldiers are exclusively dedicated to defense. First, we investigated whether defensive traits are under selection in sterile termite soldiers using allometric analyses. In soldiers of the genus Cryptotermes phragmotic traits such as a sculptured and foreshortened head evolve rapidly but were also lost twice. Second, we compared the scaling relationships of these weapons with those in solitary insects facing a trade-off between weapons and gonads. Defensive traits consistently had lower slopes than nondefensive traits which supports the existence of stabilizing selection on soldier phragmotic traits in order to plug galleries. Moreover, soldier head widths were colony specific and correlated with the minimum gallery diameter of a colony. This can proximately be explained by soldiers developing from different instars. The scaling relationships of these termite soldiers contrast strikingly with those of weapons of solitary insects, which are generally exaggerated (i.e., overscaling) male traits. These differences may provide important insights into trait evolution. Trade-offs constraining the development of individuals may have been uncoupled in termites by evolving different castes, each specialized for one function. When individuals in social insect are "released" from developmental constraints through the evolution of castes, this certainly contributed to the ecological and evolutionary success of social insects.     

Endocrine control of cuticular hydrocarbon profiles during worker-to-soldier differentiation in the termite Reticulitermes flavipes

The social organization of termites, unlike that of other social insects, is characterized by a highly plastic caste system. With the exception of the alates, all other individuals in a colony remain at an immature stage of development. Workers in particular remain developmentally flexible; they can switch castes to become soldiers or neotenics. Juvenile hormone (JH) is known to play a key role in turning workers into soldiers. In this study, we analyzed differences in cuticular hydrocarbon (CHC) profiles among castes, paying particular attention to the transition of workers to soldiers, in the subterranean termite species Reticulitermes flavipes. CHCs have a fundamental function in social insects as they serve as cues in inter- and intraspecific recognition. We showed that (1) the CHC profiles of the different castes (workers, soldiers, nymphs and neotenics) are different and (2) when workers were experimentally exposed to a JH analog and thus induced to become soldiers, their CHC profiles were modified before and after the worker-presoldier molt and before and after the presoldier-soldier molt.     

Characterization of termite lipophorin and its involvement in hydrocarbon transport

The transport of lipids constitutes a vital function in insects and requires the plasma lipoprotein lipophorin. In all insects examined to date, cuticular hydrocarbons are also transported through the hemolymph by lipophorin, and in social insects they play important roles not only in water proofing the cuticle but also in nestmate recognition. High-density lipophorin (HDLp), isolated from Reticulitermes flavipes plasma by KBr gradient ultracentrifugation, contains 66.2% protein and 33.8% lipids; hydrocarbons constitute its major neutral lipid (20.4% of total lipids). Anti-lipophorin serum was generated in rabbit and its specific association with lipophorin, and not with any other plasma proteins, was verified with Western blotting. Immunoprecipitation also confirmed that this antibody specifically recognizes lipophorin, because all hemolymph hydrocarbons of the termites R. flavipes and R. lucifugus and the cockroach Supella longipalpa, which associate only with lipophorin, were recovered in the immunoprecipitated protein. Cross-reactivity of the antiserum with lipophorin from related species was investigated by double immunodiffusion with 10 termite species in the genera Reticulitermes, Coptotermes, Zootermopsis, and Kalotermes, and with five cockroach species. Involvement of lipophorin in hydrocarbon transport was shown by injecting HDLp antiserum into Zootermopsis nevadensis and then monitoring the de novo biosynthesis of hydrocarbons and their transport to the cuticular surface; the antiserum significantly disrupted hydrocarbon transport. ELISA revealed a gradual increase in the lipophorin titer in successively larger R. flavipes workers, and differences among castes in lipophorin titers were highest between nymphs and first instar larvae.     

Termite queens have disproportionately more DNA in their fat body cells: reproductive division of labor and endoreduplication

Increases in DNA content caused by endoreduplication are widely observed in the metabolically active tissues of plants and animals. During egg production, insect females synthesize very large amounts of vitellogenin in their fat bodies, and female fat bodies of some insects become polyploid to accelerate vitellogenin production. Social insects have developed reproductive division of labor, wherein queens lay most of the eggs while other individuals have reduced fertility and undertake tasks required for maintaining the colony. Therefore, only queens are engaged in vitellogenin synthesis for egg production in social insects. Here, we show that termite queens have disproportionately more DNA in their fat body cells. Our DNA content analysis using flow cytometry demonstrated that more cells contained 4C‐DNA than 2C‐DNA in the fat bodies of Reticulitermes speratus queens. This high level of endoreduplication was not found in the fat body cells of other castes. This caste‐dependent doubling of DNA content in fat body cells suggests that termites exploit endoreduplication to boost egg production, in conjunction with the development of reproductive division of labor. This study highlights nuclear polyploidization as an adaptive strategy in social insects.     

Cuticular permeability of two species of Coptotermes Wasmann (Isoptera: Rhinotermitidae)

Cuticular permeability (CP) values of worker and soldier castes of Coptotermes formosanus Shiraki and Coptotermes vastator Light were determined using gravimetric techniques. Comparisons were made between castes, species, and between colonies of C. formosanus and C. vastator. CP values did not differ among either species or colony, however they did differ between castes. Data reported here and that of established data in the literature for C. formosanus are inconsistent, with the current report having values as small as half that of previously published data. Examining mean CP data by colony indicated that there is a range for Coptotermes spp. termites, from 6.84+/-1.2 to 25.61+/-1.49 microg H(2)Ocm(-2)mmHg(-1)h(-1) dependent upon caste. A novel method of examining intercaste CP variability is the use of worker:soldier CP ratios. Ratios for C. formosanus were consistent between the current study and previous reports. Mean percentage total body water (%TBW) lost at the time of death was 33.75+/-1.15% (soldiers) and 54.24+/-1.43% (workers) for C. formosanus, and 37.69+/-3.31% (soldiers) and 52.87+/-3.65% (workers) for C. vastator. Consistently, %TBW (and fresh mass) was greater in worker than in soldier castes. These data suggest the use of water storage mechanisms in worker termites, as shown for other rhinotermitids.     

扩头蔡白蚁消化与生殖系统解剖及其与黑翅土白蚁的比较研究

扩头蔡白蚁Tsaitermes ampliceps(Wang&Li)和黑翅土白蚁Odotoermes formosanus(Shiraki)是分布广泛的食木性昆虫,群居生活,具有复杂的品级分化.本试验研究了低等白蚁扩头蔡白蚁不同品级的消化和生殖系统结构,并与高等白蚁黑翅土白蚁进行了比较.结果显示扩头蔡白蚁的消化、生殖系...     

Quantitative comparison of caste differences in honeybee hemolymph

The honeybee, Apis mellifera, is an invaluable partner in agriculture around the world both for its production of honey and, more importantly, for its role in pollination. Honeybees are largely unexplored at the molecular level despite a long and distinguished career as a model organism for understanding social behavior. Like other eusocial insects, honeybees can be divided into several castes: the queen (fertile female), workers (sterile females), and drones (males). Each caste has different energetic and metabolic requirements, and each differs in its susceptibility to pathogens, many of which have evolved to take advantage of the close social network inside a colony. Hemolymph, arthropods' equivalent to blood, distributes nutrients throughout the bee, and the immune components contained within it form one of the primary lines of defense against invading microorganisms. In this study we have applied qualitative and quantitative proteomics to gain a better understanding of honeybee hemolymph and how it varies among the castes and during development. We found large differences in hemolymph protein composition, especially between larval and adult stage bees and between male and female castes but even between adult workers and queens. We also provide experimental evidence for the expression of several unannotated honeybee genes and for the detection of biomarkers of a viral infection. Our data provide an initial molecular picture of honeybee hemolymph, to a greater depth than previous studies in other insects, and will pave the way for future biochemical studies of innate immunity in this animal.     

Morphological phylogenetics of termites (Isoptera)

Isoptera (termites) are an ecologically important order, with both a high abundance and biomass in tropical ecosystems. However, there have been few phylogenetic hypotheses for termites, and we present here the first comprehensive cladistic analysis for the group. We analysed relationships between all seven termite families, including representatives of all known feeding group, plus a number of systematically critical taxa. Termite species richness is biased towards the higher termites (Termitidae), and our taxon sampling reflects this. Our analysis was based essentially on morphological characters (96 workers, 93 soldiers) plus seven biological characters. The cladistic analysis gave four equally parsimonious trees, representing two islands of topologies. The strict consensus tree is fully resolved for the higher termites, but less so for the lower termites. Overall there is low statistical support for the suggested topology, and this can be explained by the high incongruence between the data sets (worker, soldier and biological). This study highlights the particular problems of coding morphological characters in social insects with multiple castes. Without the input of additional data sets, e.g. alates, biological, behavioural and molecular, it will not be possible to obtain a well-supported termite phylogeny.