Glands Of Insects Research Articles

Trypanosoma cruzi infection modulates secreted phospholipase A2 expression in the salivary glands of Rhodnius prolixus

Phospholipases A2 (PLA2) comprise a superfamily of enzymes that specifically catalyze hydrolysis of the ester bond at the sn-2 position of glycerophospholipids, generating lysophospholipids and fatty acids. In Rhodnius prolixus, one of the main vectors of the Chagas's disease etiologic agent Trypanosoma cruzi, it was previously shown that lysophosphatidylcholine, a bioactive lipid, found in the insect's saliva, contributes to the inhibition of platelet aggregation, and increases the production of nitric oxide, an important vasodilator. Due to its role in potentially generating LPC, here we studied the PLA2 present in the salivary glands of R. prolixus. PLA2 activity is approximately 100 times greater in the epithelium than in the contents of salivary glands. Our study reveals the role of the RpPLA2XIIA gene in the insect feeding performance and in the fatty acids composition of phospholipids extracted from the salivary glands. Knockdown of RpPLA2XIIA significantly altered the relative amounts of palmitic, palmitoleic, oleic and linoleic acids. A short-term decrease in the expression of RpPLA2III and RpPLA2XIIA in the salivary glands of R. prolixus was evident on the third day after infection by T. cruzi. Taken together, our results contribute to the understanding of the role of PLA2 in the salivary glands of hematophagous insects and show that the parasite is capable of modulating even tissues that are not colonized by it.

Mating- and oviposition-dependent changes of the spermatheca and colleterial glands in the pest termite Cryptotermes brevis (Blattaria, Isoptera, Kalotermitidae).

The spermatheca and colleterial glands of female insects are organs associated with the reproductive system, responsible for sperm storage and secretion of egg coverings, respectively. Here we compared the development, secretory activity, and chemical nature of the secretion in the spermatheca and colleterial glands of different-aged females of the drywood termite Cryptotermes brevis. We also provide the ultrastructure of these organs in alate females. These structures have been poorly investigated in termites when compared to other eusocial insects (Hymenoptera) and termite-related dictyopterans (mantises and cockroaches). The spermatheca of C. brevis comprises a cone-shaped structure, connected to the genital chamber by a short duct. The colleterial glands, in turn, are divided into anterior and posterior tubules, each showing a basal trunk, and join into a common duct. Histological and histochemical analyses showed that the secretion of proteins and polysaccharides by the spermatheca takes place before pairing, but increases as females mate and store sperm. Colleterial glands of alates showed non-synchronous secretory activity, but the synthesis of products increased in egg-laying queens, together with the epithelium height. Ultrastructure of the spermatheca and colleterial glands revealed epithelia composed of class III secretory cells. Richness of mitochondria and electron-dense secretion in the spermatheca indicates synthesis and transport of content. Presence and absence of colleterial gland secretion in different individuals may reflect variable maturation stages of the females and secretory cells. Assuming that termites are iteroparous, the development and secretion of the spermatheca and colleterial glands play a crucial role for C. brevis queens.

Effects of 20-hydroxyecdysone on UVB-induced photoaging in hairless mice

We recently reported that exposure of skin to ultraviolet B (UVB) irradiation for 2 weeks induces stress and accelerates skin aging. Interestingly, aldosterone synthase is known to be crucial in generating UVB-induced stress-related responses, suggesting that drugs that regulate its activity can be used as skin antiaging agents. Through extensive drug screening, we have identified 20-hydroxyecdysone (20E), a steroidal prohormone secreted by the prothoracic glands of insects, as a potent inhibitor of UVB-induced aging. Although 20E has been shown to exert antistress and anti-collagenase effects in vitro, its effects in vivo remain unexplored. Furthermore, the pharmacological and physiological effects of 20E on UVB-mediated photoaging are poorly understood. Therefore, in this study, we investigated the effects of 20E on aldosterone synthase and UVB-induced photoaging and skin lesions in hairless mice, focusing on the stress-related hypothalamic–pituitary–adrenal axis. We confirmed that 20E inhibited aldosterone synthase and reduced corticosterone levels. When applied to a UV-induced skin aging animal model, it ameliorated UV-induced stress and protected against the decrease in collagen levels. Importantly, when the aldosterone synthase inhibitor osilodrostat, an FDA-approved drug, was applied to the UV-induced skin aging model, the stress-reducing and antiaging effects of 20E were not observed. Thus, we conclude that 20E inhibits UVB-induced skin aging by blocking aldosterone synthase and is a potential candidate to prevent skin aging.

Proteasome β3 subunit (PSMB3) controls female reproduction by promoting ecdysteroidogenesis during sexual maturation in Bactrocera dorsalis

Steroid hormone 20-hydroxyecdysone (20E) plays critical roles in reproductive development in dipterans and several other insect species. Ecdysteroidogenesis in the glands of larval or nymphal insects and other arthropods has been extensively studied, but that in the adult gonads remains largely unknown. Here we identified a proteasome β3 subunit (PSMB3) from a highly invasive pest Bactrocera dorsalis, and found that this gene was crucial for ecdysone production during female reproduction. PSMB3 was enriched in the ovary, and it was upregulated during sexual maturation. RNAi-mediated depletion of PSMB3 resulted in retarded ovarian development and decreased fecundity. Additionally, knockdown of PSMB3 reduced 20E titer in hemolymph of B. dorsalis. Molecularly, RNA sequencing and qPCR validation revealed that PSMB3 depletion suppressed the expression of 20E biosynthetic genes in the ovary and 20E responsive genes in the ovary and fat body. Furthermore, exogenous 20E rescued the inhibition of the ovarian development caused by PSMB3 depletion. Taken together, this study provides new insights into the adult reproductive development-related biological processes controlled by PSMB3, and proposed a potential eco-friendly control strategy against this notorious agricultural pest.

Functional Morphology and Development of the Colleterial Glands in Non- and Egg-Laying Females of the Pest Termite Coptotermes gestroi (Blattaria, Isoptera, Rhinotermitidae).

Colleterial glands of female insects are accessory glands responsible for producing secretions associated with egg-laying. Within Dictyoptera, they synthesize compounds of the ootheca. However, their morphology and role in termites are poorly understood. Here, we compared the morphology, development, and secretory activity of the colleterial glands between non- and egg-laying females of the pest termite Coptotermes gestroi under light and transmission electron microscopy. We also provide the first description of these glands for Rhinotermitidae. The glands are paired, divided into anterior and posterior units, which join in a common duct via basal trunks. They are highly developed within egg-laying females, especially the posterior gland, secreting glycoproteins to lubricate the genital chamber and/or stick the eggs together. Ultrastructure revealed glandular epithelia composed of bicellular units of Class 3, whose secretory activity varied between groups and units. Posterior gland of egg-laying females showed richness of mitochondria, rough endoplasmic reticulum, and secretory vesicles, including electron-dense secretory granules, indicating synthesis and transport of contents, especially proteins. The basal trunks were enfolded by muscles, supporting their role in conducting secretion. Morphophysiological modifications occur in the colleterial glands as females mature and lay eggs, and the mechanisms underlying the secretory cycle of the glands are discussed.

Gene expression in Verson's glands of the fall armyworm suggests their role in molting and immunity.

Verson's glands are segmental pairs of dermal glands attached to the epidermis in lepidopteran larvae. They produce macromolecules during intermolt period and empty them during each molt. Morphological, histochemical, developmental, and protein analysis studies have been conducted to determine the functions of Verson's glands. However, the exact role of Verson's glands remains unclear. In our previous study, a strain of transgenic fall armyworm, Spdoptera frugiperda expressing green fluorescence protein (GFP) and Systemic RNA interference defective protein 1 (SID1) from Caenorhabditis elegans was established to improve RNA interference (RNAi) efficiency. Unexpectedly, we found that GFP fluorescence was significantly brighter in Verson's glands than in other tissues. Also, RNAi efficiency improved more in Verson's glands than in other tissues. We took advantage of improved RNAi efficiency to explore the function of Verson's glands. RNA-seq analysis revealed that genes highly expressed in Verson's glands code for cuticular proteins, molting fluid proteins, hemolymph proteins, and antimicrobial peptides. Injection of dsRNA targeting essential genes, inhibitor of apoptosis (IAP), Actin, and vacuolar-type ATPase (VATPase) interfered with Verson's glands growth. These results revealed that Verson's glands may contribute to hemolymph, cuticle, molting fluid, and immune response during molting. This study also provide useful tools for future research in identifying the physiological role of Verson's glands in lepidopteran insects.

Competence of mosquitoes <i>Culex pipiens f.</i> molestus as carriers of West Nile virus under various temperature conditions

Introduction. The Culex pipiens mosquito is one of the proven vectors of the West Nile virus (WNV). Culex pipiens f. molestus (Cx. p. f. molestus) is a synanthropic, autogenous, widespread form of the species that can feed on a broad range of hosts, including humans. The temperature of the habitat of insects affects the potential for virus transmission, which determines the likelihood of them carrying the pathogen of West Nile fever.
 The goal is an experimental study of the temperature of the habitat of larvae on the competence of mosquitoes Cx. p. f. molestus as carriers of WNV.
 Materials and methods. We used a strain of the WNV (WNV_Volg601/18 genotype 2) and a laboratory culture of mosquitoes Cx. p. f. molestus. The concentration of the virus was detected by plaque formation using Vero cells. Insects were infected orally at the larval stage, with subsequent incubation at 20, 22 or 28C. 72 hours after the emergence of all adults from the pupae, the mosquitoes were immobilized by cold, the sex of imago was determined, the salivary glands were isolated from the females, and the presence of WNV in glandes and its titer were detected.
 Results. The titer of WNV sufficient to transmit the pathogen through the insect biting was observed in the salivary glands of insects kept at a temperature of 22 and 28C, with the virus titer rising with the temperature increasing. No virus was detected in the salivary glands of female insects kept at a temperature of 20C.
 Conclusion. Thus, it appears that the habitat temperature is an important factor limiting the replication and content of WNV in the salivary glands of Cx. p. f. molestus.

Up-Regulated Salivary Proteins of Brown Marmorated Stink Bug Halyomorpha halys on Plant Growth-Promoting Rhizobacteria-Treated Plants.

Plant Growth-Promoting Rhizobacteria (PGPR) induce systemic resistance (SR) in plants, decreasing the development of phytopathogens. The FZB42 strain of Bacillus velezensis is known to induce an SR against pathogens in various plant species. Previous studies suggested that it could also influence the interactions between plants and associated pests. However, insects have developed several strategies to counteract plant defenses, including salivary proteins that allow the insect escaping detection, manipulating defensive pathways to its advantage, deactivating early signaling processes, or detoxifying secondary metabolites. Because Brown Marmorated Stink Bug (BMSB) Halyomorpha halys is highly invasive and polyphagous, we hypothesized that it could detect the PGPR-induced systemic defenses in the plant, and efficiently adapt its salivary compounds to counteract them. Therefore, we inoculated a beneficial rhizobacterium on Vicia faba roots and soil, previous to plant infestation with BMSB. Salivary gland proteome of BMSB was analyzed by LC-MS/MS and a label-free quantitative proteomic method. Among the differentially expressed proteins, most were up-regulated in salivary glands of insects exposed to PGPR-treated plants for 24h. We could confirm that BMSB was confronted with a stress during feeding on PGPR-treated plants. The to-be-confirmeddefensive state of the plant would have been rapidly detected by the invasive H. halys pest, whichconsequently modified its salivary proteins. Among the up-regulated proteins, many could be associated with a role in plant defense counteraction, and more especially in allelochemicals detoxification or sequestration.

Regulation of male fertility and accessory gland gene expression by the Drosophila HR39 nuclear receptor

Successful reproduction is dependent on the transfer of male seminal proteins to females upon mating. These proteins arise from secretory tissues in the male reproductive tract, including the prostate and seminal vesicles in mammals and the accessory gland in insects. Although detailed functional studies have provided important insights into the mechanisms by which accessory gland proteins support reproduction, much less is known about the molecular mechanisms that regulate their expression within this tissue. Here we show that the Drosophila HR39 nuclear receptor is required for the proper expression of most genes that encode male accessory gland proteins. Consistent with this role, HR39 mutant males are infertile. In addition, tissue-specific RNAi and genetic rescue experiments indicate that HR39 acts within the accessory glands to regulate gene expression and male fertility. These results provide new directions for characterizing the mammalian orthologs of HR39, the SF-1 and LRH-1 nuclear receptors, both of which are required for glandular secretions and reproduction. In addition, our studies provide a molecular mechanism to explain how the accessory glands can maintain the abundant levels of seminal fluid production required to support fertility.

The Effect of the Hypertrophy Virus (MdSGHV) on the Ultrastructure of the Salivary Glands of Musca domestica (Diptera: Muscidae).

The salivary glands of insects play a key role in the replication cycle and vectoring of viral pathogens. Consequently, Musca domestica (L.) (Diptera: Muscidae) and the Salivary Gland Hypertrophy Virus (MdSGHV) serve as a model to study insect vectoring of viruses. A better understanding of the structural changes of the salivary glands by the virus will help obtain a better picture of the pathological impact the virus has on adult flies. The salivary glands are a primary route for viruses to enter a new host. As such, studying the viral effect on the salivary glands is particularly important and can provide insights for the development of strategies to control the transmission of vector-borne diseases, such as dengue, malaria, Zika, and chikungunya virus. Using scanning and transmission electron microscopic techniques, researchers have shown the effects of infection by MdSGHV on the salivary glands; however, the exact location where the infection was found is unclear. For this reason, this study did a close examination of the effects of the hypertrophy virus on the salivary glands to locate the specific sites of infection. Here, we report that hypertrophy is present mainly in the secretory region, while other regions appeared unaffected. Moreover, there is a disruption of the cuticular, chitinous lining that separates the secretory cells from the lumen of the internal duct, and the disturbance of this lining makes it possible for the virus to enter the lumen. Thus, we report that the chitinous lining acts as an exit barrier of the salivary gland.

Proteolytic enzymes in the salivary glands of the Neotropical brown stink bugEuschistus heros: Reduced activities in imidacloprid‐resistant strains

AbstractProteases present in the salivary glands of phytosuccivorous insects are responsible for the initial breakdown of dietary protein into the amino acids necessary for insect growth and development. Here, we characterised the total proteases and trypsin‐like activities in the salivary glands of the Neotropical brown stink bug,Euschistus heros(F.) and used in silico and biochemical approaches to determine the interactions between trypsin‐like enzymes and the soybean Kunitz trypsin inhibitor (SKTI). Additionally, we evaluated whether these enzyme activities were affected in individuals resistant to the actions of the insecticide imidacloprid. We used an originally field‐collected strain to select individuals resistant to imidacloprid alone (ImiGoiasRes) or synergised with piperonyl butoxide (ImiPBORes). A laboratory‐susceptible (ImiSusc) strain was also used to select piperonyl butoxide synergised imidacloprid‐resistant (ImiPBOSel) individuals. Our results revealed that pH and temperature significantly affected the specific trypsin‐like activities in imidacloprid‐susceptible individuals. Significant alterations in the total protease activities were recorded for field‐ (ImiGoiasRes) and laboratory‐selected imidacloprid‐resistant individuals (ImiPBOSelandImiPBORes). Our computational docking analysis demonstrated that the molecular interactions and the presence of the SKTI inhibitor (0.5 mg mL−1) could reduce up to 40% of the trypsin‐like activities of imidacloprid‐susceptible individuals. However, all imidacloprid‐resistant strains exhibited significantly less trypsin‐like activities and exhibited lower hydrolysing velocity (VMAX) and affinity (KM) rates, and the prior exposure to SKTI only synergised the imidacloprid toxicity for individuals ofImiFieldResstrain. Collectively, our findings demonstrate, for the first time, insecticide resistance detrimentally affected the activities of proteolytic enzymes in phytosuccivorous stink bug pests.

Morphological and Developmental Traits of the Binucleation of Male Accessory Gland Cells in the Benthic Water Bug, Aphelocheirus vittatus (Hemiptera: Aphelochiridae).

The male accessory glands (MAGs) in insects are pair(s) of internal reproductive organs that produce and secrete the plasma component of seminal fluid. In various insects, MAG size is important for male reproductive success because the fluid provides physiologically active substances and/or nutrients to females to control sperm as well as female reproductive behaviors. Although the MAG epithelial cells in most insect species are standard mononucleate cells, those in some insect taxa are binucleate due to incomplete cytokinesis (e.g., Drosophila [Fallén] [Diptera: Drosophilidae]) or cell fusion (e.g., Cimex [Linnaeus] [Hemiptera: Cimicidae]). In the case of Drosophila, the apicobasal position of the two nuclei relative to the epithelial plane changes from vertical to horizontal after nutrient intake, which allows the volume of the MAG cavity to expand effectively. On the other hand, in the case of Cimex, the positions of the two nuclei do not change apicobasally in response to feeding, but their position relative to the proximodistal axis varies depending on the tubular/spherical organ morphology. Here, we report that the MAG of the benthic water bug Aphelocheirus vittatus (Matsumura) (Hemiptera: Aphelochiridae) shows binucleation in all epithelial cells. Despite the phylogenetically close relationship between Aphelocheirus and Cimex, the MAG cells in Aphelocheirus showed a Drosophila-like apicobasal change in the position of the two nuclei in response to feeding. Furthermore, the cytological processes during binucleation are more similar to those in Drosophila (incomplete cytokinesis) than to those in Cimex (cell fusion). These results indicate that the physiological role and mechanism of binucleation in MAG cells changed during the evolution of Hemiptera.

The Effect of the Virus (MdSGHV) on the Ultrastructure of the Salivary Glands of Musca domestica L

The salivary glands of insects play a key role in the replication cycle and vectoring of viral pathogens. Consequently, Musca domestica L, and the Salivary Gland Hypertrophy Virus (MdSGHV) serve as a model to study insect vectoring of viruses. A better understanding of the structural aspects of this virus, and the tissues it invades, will help obtain a better picture of the pathological impact that the virus has on adult flies. Furthermore, since the virus inhibits mating and egg development in the adult fly, its study can lead to development of novel techniques to control this world‐wide pest for food safety improvement. Using scanning and transmission electron microscopic techniques (SEM, TEM), researchers have shown the effects of infection by MdSGHV on the salivary glands; however, the exact location among the glands where the infection was found is unclear (i.e., given that the salivary glands extend throughout the entire adult body). For this reason, this study intends to do a comprehensive examination of the ultrastructural effects of the hypertrophy virus on the salivary glands. Here, we report the clear hypertrophy present throughout the salivary glands in their entirety, as well as enveloped virus particles in the cytoplasm of infected cells. Moreover, the virus provokes an erosion of the cuticular lining that separates the epithelial cells from the lumen of the internal duct.

Sterol Hormone 20E Regulates Autophagy via Multiple Pathways in Insect

Sterol hormone 20-hydroxyecdysone (20E), specifically biosynthesized from cholesterol under the catalysis of a series of cytochrome P450 enzymes, is pivotal for insect molting and metamorphosis. Ecdysone, the precursor of 20E, is synthesized and secreted by a pair of prothoracic glands in holometabolous insects, and then is released into the hemolymph and converted to the active form 20E in peripheral tissues such as fat body, midgut and malpighian tubes during larval stage [1].

Accessory gland proteome of male tobacco cutworm, Spodoptera litura (F.) - An approach to identify proteins influencing reproductive physiology and behaviour

Abstract Male-derived accessory glands (MAGs) in insects typically transfer a mixture of proteins and other molecules during mating that have profound effect on female physiology and reproductive behaviour. In the present study, we sought to identify MAG proteins of the serious agricultural pest Spodoptera litura as they have been extensively studied in Drosophila and other insects. A total of 566 distinct proteins across gels of virgin and mated males, as well as 91 differentially expressed proteins in mated contrast to virgin males of S. litura were detected by 2D gel electrophoresis. Here, we successfully identified 20 MAG proteins that appeared in virgin males and depleted in mated males by mass spectrometry, thus predicting the transfer of proteins from males to females during mating. We analyzed the proteins for homology to other proteins with known functions available in Swiss- Prot through Mascot search engine. The proteins were categorized into six diverse functional groups: Metabolism, protein folding, antioxidation, olfaction, DNA binding/replication and cellular activity/response. Most of the MAG proteins fall into the same functional class as in other organisms, although some appear to rapidly evolve. The probable roles of the identified proteins associated with physiology and reproduction have been discussed. Thus, in this study we establish a strong precedent for future research on the potential of MAG proteins to interfere with the pest's own mechanisms resulting in its failure to reproduce successfully.

Binucleation of Accessory Gland Lobe Contributes to Effective Ejection of Seminal Fluid in Drosophila melanogaster.

The adult male accessory gland in insects is an internal reproductive organ analogous to the mammalian prostate, and secretes various components in the seminal fluid. Products of the accessory gland in the fruit fly Drosophila melanogaster are known to control reproductive behaviors in mated females, such as food uptake, oviposition rate, and rejection of re-mating with other males, all of which increase male reproductive capacity. Production of larger amounts of accessory gland products is thus thought to result in higher male reproductive success. The epithelium of the Drosophila accessory gland lobe is composed of a unique population of binucleate cells. We previously predicted, based on measurements of cell size in mono/binucleate mosaic accessory glands, that binucleation results in a higher plasticity in cell shape, enabling more effective ejection of seminal fluid. However, the actual effect of binucleation on ejection of seminal fluid or reproductive capacity remained unclear, as we were unable to generate an organ with uniformly mononucleate cells. In the present study, we generated organs in which most of the epithelial cells are mononucleate by manipulating aurora B or fizzy-related to block binucleation. Mononucleation resulted in a less elastic accessory gland lobe, which decreased ejection volume and the oviposition of mated females; these effects were particularly pronounced over the long term. These results suggest that binucleation in accessory gland epithelial cells contributes to higher plasticity in the volume of this organ, and enhances male reproductive success through enabling ejection of larger amounts of seminal fluid.

Seminal fluid protein genes of the brown planthopper, Nilaparvata lugens.

BackgroundSeminal fluid proteins (SFPs) are produced mainly in the accessory gland of male insects and transferred to females during mating, in which they induce numerous physiological and post-mating behavioral changes. The brown plant hopper (BPH), Nilaparvata lugens, is an economically important hemipterous pest of rice. The behavior and physiology of the female of this species is significantly altered by mating. SFPs in hemipteran species are still unclear.ResultsWe applied high-throughput mass spectrometry proteomic analyses to characterize the SFP composition in N. lugens. We identified 94 putative secreted SFPs, and the expression levels of these proteins was determined from the male accessory gland digital gene expression database. The 94 predicted SFPs showed high expression in the male accessory gland. Comparing N. lugens and other insect SFPs, the apparent expansion of N. lugens seminal fluid trypsins and carboxylesterases was observed. The number of N. lugens seminal fluid trypsins (20) was at least twice that in other insects. We detected 6 seminal fluid carboxylesterases in N. lugens seminal fluid, while seminal fluid carboxylesterases were rarely detected in other insects. Otherwise, new insect SFPs, including mesencephalic astrocyte–derived neurotrophic factor, selenoprotein, EGF (epidermal growth factor) domain–containing proteins and a neuropeptide ion transport-like peptide were identified.ConclusionThis work represents the first characterization of putative SFPs in a hemipeteran species. Our results provide a foundation for future studies to investigate the functions of SFPs in N. lugens and are an important addition to the available data for comparative studies of SFPs in insects.Electronic supplementary materialThe online version of this article (doi:10.1186/s12864-016-3013-7) contains supplementary material, which is available to authorized users.

Dehiscent organs used for defensive behavior of kamikaze termites of the genus Ruptitermes (Termitidae, Apicotermitinae) are not glands

During Isoptera evolution, the caste of soldiers disappeared in some Apicotermitinae termites as in the Neotropical Ruptitermes. Paired dorsolateral structures located between the metathorax and abdomen of foraging workers of Ruptitermes were previously denominated dehiscent glands, and are responsible for releasing an adhesive secretion that immobilizes enemies, causing their death. In this study, we investigated the morphology of dehiscent organs of workers of Ruptitermes reconditus, Ruptitermes xanthochiton, and Ruptitermes pitan and also second instar larvae of R. reconditus using light, laser scanning confocal, and transmission electron microscopy. Additionally, we performed a preliminary protein analysis using SDS-PAGE to further characterize the secretion of Ruptitermes dehiscent organs. Our results showed that the dehiscent organs do not exhibit the typical characteristics of the exocrine glandular cells class I, II or III of insects, suggesting that they constitute a new type of defensive organ. Thus, the denomination dehiscent gland was not used but dehiscent organ. Dehiscent organs in larvae are formed by fat body cells. In workers, dehiscent organs are composed by compact masses of cells that accumulate a defensive secretion and are poor in organelles related to the production of secretion. Since the dehiscent organs are not glands, we hypothesize that the dehiscent organs originate from larval fat body. The defensive secretion may have been produced at younger developmental stages of worker or the defensive compounds were absorbed from food and accumulated in the worker fat body. Histochemical techniques and SDS-PAGE revealed that the secretion of Ruptitermes dehiscent organs is constituted mainly by a protein of high molecular weight (200kDa). In conclusion, the dehiscent organs are extremely different from the exocrine glands of termites and other insects described until now. In fact, they seem to be a specialized fat body that is peculiar and exclusive of Ruptitermes termites.

Neurons of self-defence: neuronal innervation of the exocrine defence glands in stick insects.

BackgroundStick insects (Phasmatodea) use repellent chemical substances (allomones) for defence which are released from so-called defence glands in the prothorax. These glands differ in size between species, and are under neuronal control from the CNS. The detailed neural innervation and possible differences between species are not studied so far. Using axonal tracing, the neuronal innervation is investigated comparing four species. The aim is to document the complexity of defence gland innervation in peripheral nerves and central motoneurons in stick insects.ResultsIn the species studied here, the defence gland is innervated by the intersegmental nerve complex (ISN) which is formed by three nerves from the prothoracic (T1) and suboesophageal ganglion (SOG), as well as a distinct suboesophageal nerve (Nervus anterior of the suboesophageal ganglion). In Carausius morosus and Sipyloidea sipylus, axonal tracing confirmed an innervation of the defence glands by this N. anterior SOG as well as N. anterior T1 and N. posterior SOG from the intersegmental nerve complex. In Peruphasma schultei, which has rather large defence glands, only the innervation by the N. anterior SOG was documented by axonal tracing. In the central nervous system of all species, 3-4 neuron types are identified by axonal tracing which send axons in the N. anterior SOG likely innervating the defence gland as well as adjacent muscles. These neurons are mainly suboesophageal neurons with one intersegmental neuron located in the prothoracic ganglion. The neuron types are conserved in the species studied, but the combination of neuron types is not identical. In addition, the central nervous system in S. sipylus contains one suboesophageal and one prothoracic neuron type with axons in the intersegmental nerve complex contacting the defence gland.ConclusionsAxonal tracing shows a very complex innervation pattern of the defence glands of Phasmatodea which contains different neurons in different nerves from two adjacent body segments. The gland size correlates to the size of a neuron soma in the suboesophageal ganglion, which likely controls gland contraction. In P. schultei, the innervation pattern appears simplified to the anterior suboesophageal nerve. Hence, some evolutionary changes are notable in a conserved neuronal network.Electronic supplementary materialThe online version of this article (doi:10.1186/s12983-015-0122-0) contains supplementary material, which is available to authorized users.

A cecropin-like antimicrobial peptide with anti-inflammatory activity from the black fly salivary glands

BackgroundSeveral antimicrobial peptides (AMPs) belonging to the cecropin family have been identified from the salivary glands of different black fly species, however, the immunological functions for these molecules were poorly understood.MethodsA novel cecropin-like antimicrobial peptide (SibaCec) was purified using reverse phase high-performance liquid chromatography (RP-HPLC) from the salivary glands of the black fly Simulium bannaense. The amino acid sequence of SibaCec was determined by a combination method of automated Edman degradation and cDNA sequencing. The morphologic changes of Gram-negative bacteria Escherichia coli treated with SibaCec were assessed by scanning electron microscopy (SEM). Quantitative PCR (qPCR) was performed to analyze the mRNA expression of the inducible NO synthase (iNOS) and pro-inflammatory cytokines. Nitric oxide (NO) generation was examined using a Griess assay and the secretion of pro-inflammatory cytokines was determined by an enzyme-linked immunosorbent assay (ELISA). The activation of extracellular signal-regulated kinase (ERK), p38, and the nuclear translocation of nuclear factor-kappaB (NF-κB) were assessed by Western blotting analysis. Circular dichroism (CD) spectroscopy was performed to evaluate the secondary structure of SibaCec in solvent environment. Interaction of SibaCec with lipopolysaccharide (LPS) was studied using fluorescein isothiocyanate (FITC)- conjugated LPS aggregates. Neutralization of LPS by SibaCec was assayed with the chromogenic limulus amebocyte lysate (LAL) test. qPCR was also used to analyze the expression of SibaCec mRNA in the salivary glands of insects after oral infection with the bacteria E.coli.ResultsSibaCec possessed potent antimicrobial activity against Gram-negative bacteria, and showed low cytotoxicity toward mammalian cells. SEM analysis indicated that SibaCec killed bacteria through the disruption of cell membrane integrity. Furthermore, SibaCec significantly inhibited lipopolysaccharide (LPS)-induced production of NO and pro-inflammatory cytokines such as tumor necrosis factor-α (TNF-α), interferon-1β (IL-1β) and interferon-6 (IL-6) by blocking the activation of MAPKs and NF-κB signaling pathways. It mainly adopted an α-helix conformation in membrane-mimetic environments. SibaCec could interact and neutralize LPS. Infection of black flies with bacteria caused an upregulation of the expression of SibaCec.ConclusionsThese results demonstrated that in addition to the bactericidal capacity, SibaCec can function as immune regulator, inhibiting host secretion of inflammatory factors.Electronic supplementary materialThe online version of this article (doi:10.1186/s13071-015-1176-8) contains supplementary material, which is available to authorized users.