Effect of Short-Term Desiccation, Recovery Time, and CAPA–PVK Neuropeptide on the Immune System of the Burying Beetle Nicrophorus vespilloides

Environmental conditions, especially related to winter, are crucial for shaping activity of insect immune system. However, our previous research clearly indicates differences in the immune system functioning when the cold stress was induced in the laboratory conditions and when the beetles were collected from natural environment during winter. This is probably related to the multiplication of observed effects by simultaneous presence of different stress factors characteristic of winter, including desiccation. For these reasons, our next step was analysis of the effects of short-term desiccation and recovery time on the functioning of immune system of burying beetle Nicrophorus vespilloides. Also, the effect of Tenmo–PVK-2 (tenebrionid periviscerokinin), member of the CAPA–PVK neuropeptide family, was investigated to better understand observed changes. Short-term desiccation decreases the phagocytic activity of burying beetle haemocytes, which is correlated with a reduction in their adhesive ability. On the other hand, there was a significant increase in phenoloxidase (PO) activity and the level of proPO expression, which may suggest sealing the cuticula by melanin deposition and prevention of water loss. Additionally, the elevated level of defensin expression may be associated with the cross-talk between mechanisms, which participate in insect response to environmental stress, including pathogen infection. After 1 h of recovery time, the activity of tested cellular and humoral mechanisms was mostly back to the control level. However, inhibition of the activity of PO and down-regulation of proPO were noted. These results also indicate importance of melanin deposition during water loss. Moreover, it suggests that some changes in immune system functioning during stress conditions do not have an immune function. Interestingly, part of the effects characteristic of recovery time were also observed after the application of Tenmo–PVK-2, mainly related to haemocyte morphology. These results indicate that CAPA–PVK neuropeptides may also influence on activity of burying beetle immune system. It should be also highlighted that, because of the study of the effects of CAPA–PVK neuropeptides, homologs of vertebrate neuromedin U, the results may be interesting for search evolutionary similarities in the functioning of the neuroendocrine system of insects and vertebrates

Fragmentation study of major spirosolane-type glycoalkaloids by Collision Induced Dissociation Linear Ion Trap (CID-LIT) and Infrared Multiphoton Dissociation Fourier Transform Ion Cyclotron Resonance (IRMPD-FT-ICR) mass spectrometry

RATIONALE: Glycoalkaloids play a key role in the plant protection system against phytopathogens including fungi, viruses, bacteria, insects and worms. They can be toxic to humans if consumed in high concentrations causing gastrointestinal disturbances. METHODS: The structural characterization of the major spirosolane glycoalkaloids, solasonine, solamargine, -tomatine and dehydrotomatine, were investigated by positive electrospray ionization (ESI) coupled with a hybrid linear ion trap (LIT) and Fouriertransform ion cyclotron resonance (FTICR) mass spectrometer. Tandem mass spectrometric analysis of spirosolane glycoalkaloids was performed by both collision-induced dissociation (CID) within the linear ion trap and infrared multiphoton dissociation (IRMPD) in conjunction with the FTICR cell. RESULTS: Several common product ions were observed, generated by losses of sugar moiety or aglycon fragmentation in B- or E-rings, that can provide information on the accurate mass of aglycon and the primary sequence and branching of the oligosaccharide chains. Thanks to the multistage CID it was possible to understand the fragmentation pathways and thanks to high resolution of IRMPD-FTICR the elemental composition of product ions was obtained. CONCLUSIONS: Because the investigated tandem mass spectra data were acquired with high mass accuracy, unambiguous interpretation and determination of the chemical composition for the majority of detected fragment ions were feasible. From these data, generalized fragmentation pathways were proposed, providing guidance for the characterization of unknown glycoalkaloids in plants

The role of the agonists and antagonists of the muscarinic cholinergic receptors in the regulation of physiological processes in the beetle Tenebrio molitor L.

Wydział BiologiiPrzeprowadzone badania aktywności fizjologicznej agonistów (karbacholu, pilokarpiny) i antagonistów (atropiny, skopolaminy) receptorów muskarynowych pozwoliły na ocenę udziału tych receptorów w regulacji funkcjonowania mięśni narządów trzewnych, zawartości składników energetycznych w tkankach, homeostazy wolnych cukrów w hemolimfie oraz morfologii, zdolności adhezyjnych i żywotności hemocytów chrząszcza Tenebrio molitor. W pracy wykazano narządowo-specyficzne działania miotropowe badanych związków na aktywność kurczliwą serca, jelita i jajowodu. Uzyskane dane wskazują na właściwości inhibicyjne antagonistów względem serca oraz stymulujący, dawko-zależny wpływ na kurczliwość jelita i jajowodu in vitro. Biotesty in vivo i in vitro dowodzą udziału badanych związków w regulacji przemian węglowodanów i lipidów, a ich działanie jest tkankowo-specyficzne i czasowo-zależne. W badaniach wykazano, iż karbachol i skopolamina mają działanie hemocytotropowe, wpływając na morfologię, adhezję i żywotność hemocytów. Związki te powodowały apoptozę hemocytów. Immunodetekcja mAChRs w jelicie sugeruje, że działanie badanych związków na fizjologię jelita może wynikać z bezpośredniego ich wpływu na mAChRs, a identyfikacja receptorów w brzusznym łańcuszku nerwowym wskazuje na pośrednie oddziaływanie tych związków na procesy fizjologiczne przez ich wpływ na struktury nerwowe unerwiające narządy trzewne.The studies on influence of agonists (carbachol, pilocarpine) and antagonists (atropine, scopolamine) of muscarinic receptors on physiological processes in Tenebrio molitor beetle showed their participation in regulation of visceral muscles contraction, energy components content in tissues, free carbohydrates level in hemolymph and hemocytes morphology and viability. Tested compounds showed the organ-specific myotropic activity on heart, hindgut and oviduct. Results indicate that antagonists possess inhibitory properties on the heart in vivo and exert a dose-depend stimulatory effect on hindgut and oviduct contractility in vitro. In vivo and in vitro biotest indicate the input of agonists and antagonists on regulation of carbohydrates and lipid levels in tissues and the activity of these compounds is tissue-specific and time-dependent. The obtained data show that carbachol and scopolamine possess hemocytotropic activity and affect the hemocytes morphology, adhesion and viability. These compounds induce apoptosis of hemocytes. mAChRs immunodetection in the intestine suggests that tested compounds influence on contractile activity and glycogen content in this organ which may be a result of direct effect on mAChRs. The identification of these receptors in the ventral nerve cord may indicate indirect effect of the tested compounds via neural terminals which innervate visceral organs

Mitochondrial metabolism and oxidative stress in tropical cockroach under fluctuating thermal regimes

International audienceThe cockroach Gromphadorinha coquereliana can survive under low temperature and extensive periods of cold stress. To assess energy management and insect adaptation in response to cold, we measured mitochondrial activity and oxidative stress in muscle and fat body tissues from G. coquereliana -under a fluctuating thermal regime (FTR, stressed at 4°C for 3-h on 3 consecutive days, with or without 24-h recovery after last exposure). Compared to our earlier work showing that a single exposure to cold significantly affects mitochondrial parameters, here, repeated exposure to cold triggered an acclimatory response, resulting in unchanged mitochondrial bioenergetics. Immediately after cold exposure, we observed an increase in the overall pool of ATP and a decrease in typical antioxidant enzyme activity. We also observed decreased activity of uncoupling protein 4 in muscle mitochondria. After 24-h of recovery, we observed an increase in antioxidant enzymes expression in muscle and fat body and a significant increase in the expression of UCP4 and HSP in the latter. This indicates that processes related to energy conversion and disturbance under cold stress may trigger different protective mechanisms in these tissues, and these mechanisms must be activated to restore insect homeostasis. In conclusion, the measured mitochondrial parameters and the enzymatic assays results suggest that mitochondria are not affected during FTR but oxidative stress markers are decreased, and a 24-h recovery period allows for the restoration of redox and energy homeostasis, especially in the fat body. This confirms the crucial role of the fat body in intermediary metabolism and energy management in insects and in the response to repeated thermal stress

Insects as a New Complex Model in Hormonal Basis of Obesity

Nowadays, one of the biggest problems in healthcare is an obesity epidemic. Consumption of cheap and low-quality energy-rich diets, low physical activity, and sedentary work favor an increase in the number of obesity cases within many populations/nations. This is a burden on society, public health, and the economy with many deleterious consequences. Thus, studies concerning this disorder are extremely needed, including searching for new, effective, and fitting models. Obesity may be related, among other factors, to disrupting adipocytes activity, disturbance of metabolic homeostasis, dysregulation of hormonal balance, cardiovascular problems, or disorders in nutrition which may lead to death. Because of the high complexity of obesity, it is not easy to find an ideal model for its studies which will be suitable for genetic and physiological analysis including specification of different compounds’ (hormones, neuropeptides) functions, as well as for signaling pathways analysis. In recent times, in search of new models for human diseases there has been more and more attention paid to insects, especially in neuro-endocrine regulation. It seems that this group of animals might also be a new model for human obesity. There are many arguments that insects are a good, multidirectional, and complex model for this disease. For example, insect models can have similar conservative signaling pathways (e.g., JAK-STAT signaling pathway), the presence of similar hormonal axis (e.g., brain–gut axis), or occurrence of structural and functional homologues between neuropeptides (e.g., neuropeptide F and human neuropeptide Y, insulin-like peptides, and human insulin) compared to humans. Here we give a hint to use insects as a model for obesity that can be used in multiple ways: as a source of genetic and peptidomic data about etiology and development correlated with obesity occurrence as well as a model for novel hormonal-based drug activity and their impact on mechanism of disease occurrence

Cholinergic Agonists and Antagonists Have an Effect on the Metabolism of the Beetle <i>Tenebrio Molitor</i>

Synthetic insecticides are still widely used in plant protection. The main target for their action is the nervous system, in which the cholinergic system plays a vital role. Currently available insecticides have low selectivity and act on the cholinergic systems of invertebrates and vertebrates. Acetylcholine, a cholinergic system neurotransmitter, acts on cells by two types of receptors: nicotinic and muscarinic. In mammals, the role of muscarinic acetylcholine receptors (mAChRs) is quite well-known but in insects, is still not enough. Based on data indicating that the muscarinic cholinergic system strongly affects mammalian metabolism, we investigated if it similarly occurs in insects. We investigated the influence of agonists (acetylcholine, carbachol, and pilocarpine) and antagonists (tropane alkaloids: atropine and scopolamine) of mAChRs on the level of selected metabolites in Tenebrio molitor beetle trophic tissues. We analyzed the glycogen content in the fat body and midgut, the total free sugar concentration in the hemolymph and the lipid amount in the fat body. Moreover, we analyzed the levels of insulin-like peptides in the hemolymph. The tested compounds significantly influenced the mentioned parameters. They increased the glycogen content in the fat body and midgut but decreased the concentration of free sugars in the hemolymph. The observed effects were tissue-specific, and were also time- and dose-dependent. We used nonligated and neck-ligated larvae (to eliminate the influence of head factors on tissue metabolism) to determine whether the observed changes are the result of direct or indirect impacts on tissues. The obtained data suggest that the cholinergic system affects the fat body and midgut indirectly and directly and a pleiotropic role for mAChRs exists in the regulation of energy metabolism in insects. Moreover, tested compounds significantly affected the level of insulin-like peptides in hemolymph. Our studies for the first time showed that mAChRs are involved in regulation of insect metabolism of trophic tissues, and act on them directly and indirectly. Improved knowledge about insect cholinergic system may help in searching more selective and environment-friendly solutions in pest management

Thermal stress causes DNA damage and mortality in a tropical insect

International audienceCold tolerance is considered an important factor determining the geographic distribution of insects. We have previously shown that despite its tropical origin, the cockroach is capable of surviving exposures to cold. However, the freezing tolerance of this species had not yet been examined. Low temperature is known to alter membrane integrity in insects, but whether chilling or freezing compromises DNA integrity remains a matter of speculation. In the present study, we subjected the adults to freezing to determine their supercooling point (SCP) and evaluated whether the cockroaches were capable of surviving partial and complete freezing. Next, we conducted single cell gel electrophoresis (SCGE) assays to determine whether heat, cold and freezing altered hemocyte DNA integrity. The SCP of this species was high and around -4.76°C, which is within the typical range of freezing-tolerant species. Most cockroaches survived to 1 day after partial ice formation (20% mortality), but died progressively in the next few days after cold stress (70% mortality after 4 days). One day after complete freezing, most insects died (70% mortality), and after 4 days, 90% of them had succumbed. The SCGE assays showed substantial levels of DNA damage in hemocytes. When cockroaches were heat-stressed, the level of DNA damage was similar to that observed in the freezing treatment, though all heat-stressed insects survived. The present study shows that can be considered as moderately freeze-tolerant, and that extreme low temperature stress can affect DNA integrity, suggesting that this cockroach may possess an efficient DNA repair system

The effects of cold stress on tropical cockroach Gromphadorhina portentosa

International audienc

The effects of cold stress on tropical cockroach Gromphadorhina portentosa

International audienc

Plant secondary metabolites as potential bioinsecticides? Study of the effects of plant-derived volatile organic compounds on the reproduction and behaviour of the pest beetle Tenebrio molitor

Modern agriculture has many environmental consequences, such as soil contamination, accumulation of toxic compounds in the environment or risk of adverse effects on nontarget organisms and for these reasons, scientists are seeking a more environmentally friendly alternative to synthetic insecticides. This study investigated the effects of four plant secondary metabolites classified as volatile organic compounds (VOCs), which have potential as bioinsecticides, (E)-2-decenal, furfural, 2-undecanone and (E,E)-2–4-decadienal, in concentrations 10−5 and 10−7 M, on female reproductive processes and larval hatchability of the Tenebrio molitor beetle. Our study indicates proper development of ovaries after application of compounds however the volume of terminal oocytes was significantly reduced, with the strongest effect of (E)− 2-decenal which reduced the volume approximately three times. The relative vitellogenin expression level was reduced, with the strongest effect observed after application of furfural, (E,E)− 2–4-decadienal and (E)− 2-decenal in concentration 10−7 M, at the same time patency index was significantly reduced up to 2-times after application of furfural at 10−7 M. What is more important morphological changes translated into physiological ones. The number of laid eggs was affected, with the strongest inhibition after application of furfural (∼43% reduction), (E,E)− 2–4-decadienal (∼33%) and (E)− 2-decenal at concentration 10−7 M (∼33%). Moreover, we observed up to 13% (in case of 2-undecanone) decrease in larval hatchability. Tested compounds exhibited a repellent effect and caused 60% reduction of insect survivability after (E)− 2-decenal at concentration 10−5 M. Altogether, VOCs seems like potential bioactive compounds in plant protection