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Imidacloprid is a well-known systemic insecticide which has a deleterious impact on honeybees. Beekeepers in the Tizi-Ouzou wilaya (Algeria) where the imidacloprid insecticide is used, report unusual losses and deaths of bee colonies. Even at sublethal doses, insecticid can impact the most crucial tasks of a bee colony such as comb building. This study was conducted in order to investigate the effect of an imidacloprid based insecticide (Confidor®Supra) on the production of wax by the honeybee Apis mellifera intermissa. After the imidacloprid LD50 was determined in controlled conditions, three sublethal doses were tested. The mortality, the syrup consumption and the weight of the wax generated were recorded. The imidacloprid insecticide LD50 at 48 hours was evaluated at 3.5 ng.per bee on 4 days old spring worker bees. We found that the three sublethal doses (0.175 mg L-1, 0.087 mg L-1 and 0.035 mg L-1) had an impact on the syrup consumption and the wax production by adult bees. Bees exposed to sublethal doses of insecticide consumed less syrup and produced less wax that the control bees. A dose response was observed regarding the production of wax. The reduction of wax production by bees caused by an exposure to an imidacloprid based insecticide ads up to the many other effects of imidacloprid described in the literature.  This kind of impact could have harmful consequences for bee colonies as wax production is the basis of nest building. The physiological causes of the reduction of wax production remain to be investigated. [ABSTRACT FROM AUTHOR]

Abstract: Aim: In vitro and in vivo studies were conducted to test a new carvacrol‐based product designed to delay the carvacrol release so that it could reach the caeca of broiler chickens in order to control Campylobacter jejuni. Methods and Results: Antimicrobial activity of carvacrol, a constituent of oregano and thyme essential oil, has been demonstrated against C. jejuni in vitro, and this compound was found beneficial for broiler growth. Here, we tested a new liquid formulation that did not change the antibacterial efficacy of carvacrol against C. jejuni in vitro, as assessed by broth microdilution. The mode of action of carvacrol also remained unchanged as illustrated by electronic microscopy. A pharmacokinetic assay monitored carvacrol of the solid galenic formulation in the avian digestive tract and this showed that this compound was mainly found in the last part (caeca, large intestine) and in the droppings. Extremely low concentrations of free carvacrol were present in blood plasma, with larger amounts of carvacrol metabolites: carvacrol glucuronide and sulphate. A qPCR analysis showed that the solid galenic form of carvacrol added at 5 kg per tonne of food (i.e. 9·5 mg of carvacrol per kg of bodyweight per day) significantly decreased the C. jejuni caecal load by 1·5 log. Conclusions: The new liquid formulation was as effective as unformulated carvacrol in vitro. In vivo the solid galenic form seems to delay the carvacrol release into the caeca and presented interesting results on C. jejuni load after 35 days. Significance and Impact of the Study: Results suggested that this product could be promising to control Campylobacter contamination of broilers. [ABSTRACT FROM AUTHOR]

Plant extracts are considered as a large source of active biomolecules, especially in phytosanitary and pharmacological fields. Anthyllis henoniana is a woody Saharan plant located in the big desert of North Africa. Our previous research paper proved the richness of the methanol extract obtained from the stems in flavonoids and phenolic compounds as well as its remarkable antioxidant activity. In this research, we started by investigating the phytochemical composition of the methanol extract using high performance liquid chromatography coupled with electrospray ionization mass spectrometry (LC-MS/MS). Among the 41 compounds identified, we isolated and characterized (structurally and functionally) the most abundant product, a flavonoid triglycoside (AA 770 ) not previously described in this species. This compound, which presents no cytotoxic activity, exhibits an interesting cellular antioxidant effect by reducing reactive oxygen species (ROS) generation, and an antiproliferative action on breast cancer cells. This study provides a preliminary investigation into the pharmacological potential of the natural compound AA 770 , isolated and identified from Anthyllis henoniana for the first time., Competing Interests: The authors declare no conflicts of interest.

The mast cell receptor Mrgprb2, a mouse orthologue of human Mrgprx2, is known as an inflammatory receptor and its elevated expression is associated with various diseases such as ulcerative colitis. We aimed to elucidate the role of Mrgprb2/x2 and the effect of its ligands on a chemically induced murine colitis model. We showed that in Mrgprb2 -/- mice, there is a differential regulation of cytokine releases in the blood plasma and severe colonic damages after DSS treatment. Unexpectedly, we demonstrated that known Mrgprb2/x2 agonists (peptide P17, P17 analogues and CST-14) and antagonist (GE1111) similarly increased the survival rate of WT mice subjected to 4% DSS-induced colitis, ameliorated the colonic damages of 2.5% DSS-induced colitis, restored major protein mRNA expression involved in colon integrity, reduced CD68 + and F4/80 + immune cell infiltration and restored cytokine levels. Collectively, our findings highlight the eminent role of Mrpgrb2/x2 in conferring a beneficial effect in the colitis model, and this significance is demonstrated by the heightened severity of colitis with altered cytokine releases and inflammatory immune cell infiltration observed in the Mrgprb2 knockout mice. Elevated expression of Mrgprb2 in WT colitis murine models may represent the organism's adaptive protective mechanism since Mrgprb2 knockout results in severe colitis. On the other hand, both agonist and antagonist of Mrgprb2 analogously mitigated the severity of colitis in DSS-induced colitis model by altering Mrgprb2 expression, immune cell infiltration and inflammatory cytokine releases., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024. Published by Elsevier Masson SAS.)

Background: Venoms have evolved independently over a hundred times in the animal kingdom to deter predators and/or subdue prey. Venoms are cocktails of various secreted toxins, whose origin and diversification provide an appealing system for evolutionary researchers. Previous studies of the ant venom of Tetramorium bicarinatum revealed several Myrmicitoxin (MYRTX) peptides that gathered into seven precursor families suggesting different evolutionary origins. Analysis of the T. bicarinatum genome enabling further genomic approaches was necessary to understand the processes underlying the evolution of these myrmicitoxins., Results: Here, we sequenced the genome of Tetramorium bicarinatum and reported the organisation of 44 venom peptide genes (vpg). Of the eleven chromosomes that make up the genome of T. bicarinatum, four carry the vpg which are organized in tandem repeats. This organisation together with the ML evolutionary analysis of vpg sequences, is consistent with evolution by local duplication of ancestral genes for each precursor family. The structure of the vpg into two or three exons is conserved after duplication events while the promoter regions are the least conserved parts of the vpg even for genes with highly identical sequences. This suggests that enhancer sequences were not involved in duplication events, but were recruited from surrounding regions. Expression level analysis revealed that most vpg are highly expressed in venom glands, although one gene or group of genes is much more highly expressed in each family. Finally, the examination of the genomic data revealed that several genes encoding transcription factors (TFs) are highly expressed in the venom glands. The search for binding sites (BS) of these TFs in the vpg promoters revealed hot spots of GATA sites in several vpg families., Conclusion: In this pioneering investigation on ant venom genes, we provide a high-quality assembly genome and the annotation of venom peptide genes that we think can fosters further genomic research to understand the evolutionary history of ant venom biochemistry., (© 2024. The Author(s).)

Ants are among the most abundant terrestrial invertebrate predators on Earth. To overwhelm their prey, they employ several remarkable behavioral, physiological, and biochemical innovations, including an effective paralytic venom. Ant venoms are thus cocktails of toxins finely tuned to disrupt the physiological systems of insect prey. They have received little attention yet hold great promise for the discovery of novel insecticidal molecules. To identify insect-neurotoxins from ant venoms, we screened the paralytic activity on blowflies of nine synthetic peptides previously characterized in the venom of Tetramorium bicarinatum . We selected peptide U 11 , a 34-amino acid peptide, for further insecticidal, structural, and pharmacological experiments. Insecticidal assays revealed that U 11 is one of the most paralytic peptides ever reported from ant venoms against blowflies and is also capable of paralyzing honeybees. An NMR spectroscopy of U 11 uncovered a unique scaffold, featuring a compact triangular ring helix structure stabilized by a single disulfide bond. Pharmacological assays using Drosophila S2 cells demonstrated that U 11 is not cytotoxic, but suggest that it may modulate potassium conductance, which structural data seem to corroborate and will be confirmed in a future extended pharmacological investigation. The results described in this paper demonstrate that ant venom is a promising reservoir for the discovery of neuroactive insecticidal peptides.

Stings of certain ant species (Hymenoptera: Formicidae) can cause intense, long-lasting nociception. Here we show that the major contributors to these symptoms are venom peptides that modulate the activity of voltage-gated sodium (Na V ) channels, reducing their voltage threshold for activation and inhibiting channel inactivation. These peptide toxins are likely vertebrate-selective, consistent with a primarily defensive function. They emerged early in the Formicidae lineage and may have been a pivotal factor in the expansion of ants., (© 2023. The Author(s).)

Anthyllis henoniana stems were harvested in two seasons: winter and spring (February and May 2021). In this study, we investigated the antioxidant (DPPH, ABTS, FRAP and TAC) and antimicrobial activities, total phenolic contents and total flavonoid contents of the obtained extracts (hexane, ethyl acetate and methanol). The results showed that ethyl acetate extract from stems harvested in winter exhibited the highest antioxidant activity, while ethyl acetate extract from the stems harvested in spring showed the most potent antibacterial and antifungal activities. To explain these differences, we investigated the phytochemical composition of these two extracts using liquid chromatography coupled with mass spectrometry. Therefore, 45 compounds were detected, from which we identified 20 compounds (flavonoids, triterpenoids, chalcones and phenolic acids); some were specific to the harvest month while others were common for both periods. Some of the major compounds detected in ethyl acetate (spring) were dihydrochalcone (Kanzonol Y, 8.2%) and flavanone (sophoraflavanone G, 5.9%), previously recognized for their antimicrobial effects. We therefore concluded that the difference in activities observed for the two harvest periods depends on the chemical composition of the extracts and the relative abundance of each compound.

Hymenopterans represent one of the most abundant groups of venomous organisms but remain little explored due to the difficult access to their venom. The development of proteo-transcriptomic allowed us to explore diversity of their toxins offering interesting perspectives to identify new biological active peptides. This study focuses on U 9 function, a linear, amphiphilic and polycationic peptide isolated from ant Tetramorium bicarinatum venom. It shares physicochemical properties with M-Tb1a, exhibiting cytotoxic effects through membrane permeabilization. In the present study, we conducted a comparative functional investigation of U 9 and M-Tb1a and explored the mechanisms underlying their cytotoxicity against insect cells. After showing that both peptides induced the formation of pores in cell membrane, we demonstrated that U 9 induced mitochondrial damage and, at high concentrations, localized into cells and induced caspase activation. This functional investigation highlighted an original mechanism of U 9 questioning on potential valorization and endogen activity in T . bicarinatum venom., Competing Interests: The authors declare no competing interests., (© 2023.)

Among ants, Myrmicinae represents the most speciose subfamily. The venom composition previously described for these social insects is extremely variable, with alkaloids predominant in some genera while, conversely, proteomics studies have revealed that some myrmicine ant venoms are peptide-rich. Using integrated transcriptomic and proteomic approaches, we characterized the venom peptidomes of six ants belonging to the different tribes of Myrmicinae. We identified a total of 79 myrmicitoxins precursors which can be classified into 38 peptide families according to their mature sequences. Myrmicine ant venom peptidomes showed heterogeneous compositions, with linear and disulfide-bonded monomers as well as dimeric toxins. Several peptide families were exclusive to a single venom whereas some were retrieved in multiple species. A hierarchical clustering analysis of precursor signal sequences led us to divide the myrmicitoxins precursors into eight families, including some that have already been described in other aculeate hymenoptera such as secapin-like peptides and voltage-gated sodium channel (Na V ) toxins. Evolutionary and structural analyses of two representatives of these families highlighted variation and conserved patterns that might be crucial to explain myrmicine venom peptide functional adaptations to biological targets., Competing Interests: Declaration of competing interest None., (Copyright © 2022. Published by Elsevier Ltd.)

The phytochemical analysis of antioxidant and antibacterial activities of Erodium arborescens aerial part extracts constitute the focus of this research. The chemical composition of an acetone extract was investigated using LC-HESI-MS 2 , which revealed the presence of 70 compounds. The major identified components were tannin derivatives. Total polyphenol and total flavonoid contents were assessed in plant extracts (hexane, ethyl acetate, acetone and methanol). The results showed that the acetone extract exhibited the highest contents of polyphenols and flavonoids, 895.54 and 36.39 mg QE/g DE, respectively. Furthermore, when compared to other extracts, Erodium arborescens acetone extract was endowed with the highest antioxidant activity with 2,2-diphenyl-1-picrylhydrazyl (DPPH), ferric reducing antioxidant power (FRAP) and total antioxidant capacity (TAC) tests. In addition, the four extracts of Erodium arborescens showed variable degrees of antimicrobial activity against the tested strains, and the interesting activity was obtained with acetone and methanol extracts.

Aim: This study was conducted to test the ability of a carvacrol-based formulation (Phodé, France) to decrease the C. jejuni caecal load in inoculated broiler chickens and to study the impact of the C. jejuni inoculation alone or combined with the product, on the caecal microbiota., Methods and Results: On day 1, chickens were either fed a control feed or the same diet supplemented with a carvacrol-based product. On day 21, the carvacrol-supplemented chickens and half of the non-supplemented chickens were inoculated with C. jejuni (10 8  CFU). Quantitative PCR was used to quantify C. jejuni in chicken caecal samples and 16S rRNA gene sequencing was carried out at 25, 31 and 35 days of age. A significant decrease of 1.4 log of the C. jejuni caecal load was observed in 35-day-old chickens supplemented with the product, compared to the inoculated and unsupplemented group (p < 0.05). The inoculation with C. jejuni significantly increased the population richness, Shannon and Simpson diversity and altered beta-diversity. Compared to the control group, the C. jejuni inoculation causes significant changes in the microbiota. The carvacrol-based product associated with C. jejuni inoculation increased the diversity and strongly modified the structure of the microbial community. Functional analysis by 16S rRNA gene-based predictions further revealed that the product up-regulated the pathways involved in the antimicrobial synthesis, which could explain its shaping effect on the caecal microbiota., Conclusions: Our study confirmed the impairment of the caecal bacterial community after inoculation and demonstrated the ability of the product to reduce the C. jejuni load in chickens. Further investigations are needed to better understand the mode of action of this product to promote the installation of a beneficial microbiota to its host., Significance and Impact of the Study: Results suggested that this product could be promising to control C. jejuni contamination of broilers., (© 2022 Phodé. Journal of Applied Microbiology published by John Wiley & Sons Ltd on behalf of Society for Applied Microbiology.)

Exposure to pesticides through eyes, skin, ingestion and inhalation may affects human health by interfering with immune cells, such as macrophages. We evaluated, in vitro, the effect of six pesticides widely used in apple arboriculture on the functions of human monocyte-derived macrophages (hMDMs). hMDMs were cultured for 4 or 24 h with or without pesticides (0.01, 0.1, 1, 10 μmol.L -1 ). We showed that chlorpyrifos, thiacloprid, thiophanate, boscalid, and captan had little toxic effect at the tested concentrations, while dithianon had low-cytotoxicity at 10 μmol.L -1 . While boscalid showed no effect on hMDMs function, thiophanate (0.01 μmol.L -1 ) stimulated with TPA and thiacloprid (1, 10 μmol.L -1 ) stimulated with zymosan activated ROS production. Chlorpyrifos, dithianon, and captan inhibited ROS production and TNF-α, IL-1β pro-inflammatory cytokines. We established that dithianon (0.01-1 μmol.L -1 ) and captan (0.1, 1 μmol.L -1 ) induced mRNA expression of NQO1 and HMOX1 antioxidant enzymes. Dithianon also induced the mRNA expression of catalase, superoxide dismutase-2 at 10 μmol.L -1 . Together, these results show that exposure to chlorpyrifos, dithianon, and captan induce immunomodulatory effects that may influence the disease fighting properties of monocytes/macrophages while pesticides such as thiacloprid, thiophanate and boscalid have little influence., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

Considering medicinal plants as an inexhaustible source of active ingredients that may be easily isolated using simple and inexpensive techniques, phytotherapy is becoming increasingly popular. Various experimental approaches and analytical methods have been used to demonstrate that the genus Calendula (Asteraceae) has a particular richness in active ingredients, especially phenolic compounds, which justifies the growing interest in scientific studies on this genus' species. From a chemical and biological viewpoint, Calendula aegyptiaca is a little-studied plant. For the first time, high-performance liquid chromatography combined with negative electrospray ionization mass spectrometry (HPLC-HESI-MS) was used to analyze methanolic extracts of Calendula aegyptiaca ( C. aegyptiaca ) fruits. Thirty-five molecules were identified. Flavonoids (47.87%), phenolic acids (5.18%), and saponins (6.47%) formed the majority of these chemicals. Rutin, caffeic acid hexoside, and Soyasaponin βg' were the most abundant molecules in the fruit methanolic extract, accounting for 17.49% of total flavonoids, 2.32 % of total phenolic acids, and 0.95% of total saponins, respectively. The antioxidant activity of the fruit extracts of C. aegyptiaca was investigated using FRAP, TAC, and DPPH as well as flavonoids and total phenols content. Because the phenolic components were more extractable using polar solvents, the antioxidant activity of the methanolic extract was found to be higher than that of the dichloromethane and hexane extracts. The IC50 value for DPPH of methanolic extract was found to be 0.041 mg·mL -1 . Our findings showed that C. aegyptiaca is an important source of physiologically active compounds.

Background: P17, a peptide isolated from Tetramorium bicarinatum ant venom, is known to induce an alternative phenotype of human monocyte-derived macrophages via activation of an unknown G protein-coupled receptor (GPCR)., Objective: We sought to investigate the mechanism of action and the immunomodulatory effects of P17 mediated through MRGPRX2 (Mas-related G protein-coupled receptor X2)., Methods: To identify the GPCR for P17, we screened 314 GPCRs. Upon identification of MRGPRX2, a battery of in silico, in vitro, ex vivo, and in vivo assays along with the receptor mutation studies were performed. In particular, to investigate the immunomodulatory actions, we used β-hexosaminidase release assay, cytokine releases, quantification of mRNA expression, cell migration and differentiation assays, immunohistochemical labeling, hematoxylin and eosin, and immunofluorescence staining., Results: P17 activated MRGPRX2 in a dose-dependent manner in β-arrestin recruitment assay. In LAD2 cells, P17 induced calcium and β-hexosaminidase release. Quercetin- and short hairpin RNA-mediated knockdown of MRGPRX2 reduced P17-evoked β-hexosaminidase release. In silico and in vitro mutagenesis studies showed that residue Lys 8 of P17 formed a cation-π interaction with the Phe 172 of MRGPRX2 and [Ala 8 ]P17 lost its activity partially. P17 activated LAD2 cells to recruit THP-1 and human monocytes in Transwell migration assay, whereas MRGPRX2-impaired LAD2 cells cannot. In addition, P17-treated LAD2 cells stimulated differentiation of THP-1 and human monocytes, as indicated by the enhanced expression of macrophage markers cluster of differentiation 11b and TNF-α by quantitative RT-PCR. Immunohistochemical and immunofluorescent staining suggested monocyte recruitment in mice ears injected with P17., Conclusions: Our data provide novel structural information regarding the interaction of P17 with MRGPRX2 and intracellular pathways for its immunomodulatory action., (Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.)

Aeonium is a genus of succulents belonging to the Crassulaceae family. Their importance in traditional medicine has stimulated both pharmacological and chemical research. In this study, we optimized extraction, separation, and analytical conditions using a high performance liquid chromatographic method coupled with electrospray ionization mass spectrometry by the negative mode (HPLC-ESI-MS) in order to, for the first time, determine thirty-four compounds from Aeonium arboreum leaves. Twenty-one of them are assigned among which are sixteen flavonoids and five phenolic acids. FRAP, TAC, DPPH, and ABTS •+ radical scavenging were used to evaluate antioxidant activity. The obtained IC 50 values ranged from 0.031 to 0.043 mg.mL -1 for DPPH and between 0.048 and 0.09 mg·mL -1 for ABTS •+ . Antimicrobial activity was also assessed. The obtained minimum inhibitory concentrations (MIC) of these extracts ranged from 12.5 to 50 µg·mL -1 against Micrococcus luteus , Listeria ivanovii , Staphylococcus aureus , Salmonella enterica , Escherichia coli , Pseudomonas aeruginosa , Aspergillus niger , and Fusarium oxysporum , and from 25 to 50 µg·mL -1 against Candida albicans . Therefore, these extracts can be considered as a potential source of biological active compounds.

Ziram, a zinc dithiocarbamate is widely used worldwide as a fungicide in agriculture. In order to investigate ziram-induced changes in macrophage functions and polarization, human monocytes-derived macrophages in culture were treated with ziram at 0.01-10 μmol.L -1 for 4-24 h. To characterize zinc involvement in these changes, we also determined the effects of disulfiram alone (dithiocarbamate without zinc) or in co-incubation with ZnSO 4 . We have shown that ziram and disulfiram at 0.01 μmol.L -1 increased zymosan phagocytosis. In contrast, ziram at 10 μmol.L -1 completely inhibited this phagocytic process, the oxidative burst triggered by zymosan and the production of TNF-α, IL-1β, IL-6, and CCL2 triggered by LPS. Disulfiram had the same effects on these macrophages functions only when combined with zinc (10 μmol.L -1 ). In contrast, at 10 μmol.L -1 ziram and zinc associated-disulfiram induced expression of several antioxidants genes HMOX1, SOD2, and catalase, which could suggest the induction of oxidative stress. This oxidative stress could be involved in the increase in late apoptosis induced by ziram (10 μmol.L -1 ) and zinc associated-disulfiram. Concerning gene expression profiles of membrane markers of macrophage polarization, ziram at 10 μmol.L -1 had two opposite effects. It inhibited the gene expression of M2 markers (CD36, CD163) in the same way as the disulfiram-zinc co-treatment. Conversely, ziram induced gene expression of other M2 markers CD209, CD11b, and CD16 in the same way as treatment with zinc alone. Disulfiram-zinc association had no significant effects on these markers. These results taken together show that ziram via zinc modulates macrophages to M2-like anti-inflammatory phenotype which is often associated with various diseases.

In the mutualisms involving certain pseudomyrmicine ants and different myrmecophytes (i.e., plants sheltering colonies of specialized "plant-ant" species in hollow structures), the ant venom contributes to the host plant biotic defenses by inducing the rapid paralysis of defoliating insects and causing intense pain to browsing mammals. Using integrated transcriptomic and proteomic approaches, we identified the venom peptidome of the plant-ant Tetraponera aethiops (Pseudomyrmecinae). The transcriptomic analysis of its venom glands revealed that 40% of the expressed contigs encoded only seven peptide precursors related to the ant venom peptides from the A-superfamily. Among the 12 peptide masses detected by liquid chromatography-mass spectrometry (LC-MS), nine mature peptide sequences were characterized and confirmed through proteomic analysis. These venom peptides, called pseudomyrmecitoxins (PSDTX), share amino acid sequence identities with myrmeciitoxins known for their dual offensive and defensive functions on both insects and mammals. Furthermore, we demonstrated through reduction/alkylation of the crude venom that four PSDTXs were homo- and heterodimeric. Thus, we provide the first insights into the defensive venom composition of the ant genus Tetraponera indicative of a streamlined peptidome.

Helicobacter pylori (Marshall & Goodwin) is a widespread human pathogen that is acquiring resistance to the antibiotics used to treat it. This increasing resistance necessitates a continued search for new antibiotics. An antibiotic source that shows promise is animals whose immune systems must adapt to living in bacteria-laden conditions by producing antibacterial peptides or small molecules. Among these animals is the black soldier fly (BSF; Hermetia illucens Linnaeus), a Diptera that colonizes decomposing organic matter. In order to find anti-H. pylori peptides in BSF, larvae were challenged with Escherichia coli (Enterobacteriales: Enterobacteriaceae). Small peptides were extracted from hemolymph and purified using solid-phase extraction, molecular weight cutoff filtration and two rounds of preparative high performance liquid chromatography (HPLC). The anti-H. pylori fraction was followed through the purification process using the inhibition zone assay in brain-heart infusion agar, while peptides from uninoculated larvae had no activity. The inhibition halo of the active sample was comparable to the action of metronidazole in the inhibition zone assay. The purified sample contained four peptides with average masses of approximately 4.2 kDa that eluted together when analyzed by HPLC-mass spectrometry. The peptides likely have similar sequences, activity, and properties. Therefore, BSF produces inducible antibacterial peptides that have in vitro activity against H. pylori, which highlights BSF's position as an important target for further bioprospecting., (© The Author(s) [2019]. Published by Oxford University Press on behalf of Entomological Society of America.)

The peptide toxins in the venoms of small invertebrates such as stinging ants have rarely been studied due to the limited amount of venom available per individual. We used a venomics strategy to identify the molecular diversity of the venom peptidome for the myrmicine ant Tetramorium bicarinatum. The methodology included (i) peptidomics, in which the venom peptides are sequenced through a de novo mass spectrometry approach or Edman degradation; (ii) transcriptomics, based on RT-PCR-cloning and DNA sequencing; and (iii) the data mining of the RNA-seq in the available transcriptome. Mass spectrometry analysis revealed about 2800 peptides in the venom. However, the de novo sequencing suggested that most of these peptides arose from processing or the artifactual fragmentations of full-length mature peptides. These peptides, called "myrmicitoxins", are produced by a limited number of genes. Thirty-seven peptide precursors were identified and classified into three superfamilies. These precursors are related to pilosulin, secapin or are new ant venom prepro-peptides. The mature myrmicitoxins display sequence homologies with antimicrobial, cytolytic and neurotoxic peptides. The venomics strategy enabled several post-translational modifications in some peptides such as O-glycosylation to be identified. This study provides novel insights into the molecular diversity and evolution of ant venoms.

The venom peptide bicarinalin, previously isolated from the ant Tetramorium bicarinatum , is an antimicrobial agent with a broad spectrum of activity. In this study, we investigate the potential of bicarinalin as a novel agent against Helicobacter pylori , which causes several gastric diseases. First, the effects of synthetic bicarinalin have been tested against Helicobacter pylori : one ATCC strain, and forty-four isolated from stomach ulcer biopsies of Peruvian patients. Then the cytoxicity of bicarinalin on human gastric cells and murine peritoneal macrophages was measured using XTT and MTT assays, respectively. Finally, the preventive effect of bicarinalin was evaluated by scanning electron microscopy using an adherence assay of H. pylori on human gastric cells treated with bicarinalin. This peptide has a potent antibacterial activity at the same magnitude as four antibiotics currently used in therapies against H. pylori . Bicarinalin also inhibited adherence of H. pylori to gastric cells with an IC 50 of 0.12 μg·mL -1 and had low toxicity for human cells. Scanning electron microscopy confirmed that bicarinalin can significantly decrease the density of H. pylori on gastric cells. We conclude that Bicarinalin is a promising compound for the development of a novel and effective anti- H. pylori agent for both curative and preventive use., Competing Interests: The authors declare no conflict of interest.

Despite the growing knowledge with regard to the immunomodulatory properties of host defense peptides, their impact on macrophage differentiation and on its associated microbicidal functions is still poorly understood. Here, we demonstrated that the P17, a new cationic antimicrobial peptide from ant venom, induces an alternative phenotype of human monocyte-derived macrophages (h-MDMs). This phenotype is characterized by a C-type lectin receptors (CLRs) signature composed of mannose receptor (MR) and Dectin-1 expression. Concomitantly, this activation is associated to an inflammatory profile characterized by reactive oxygen species (ROS), interleukin (IL)-1β, and TNF-α release. P17-activated h-MDMs exhibit an improved capacity to recognize and to engulf Candida albicans through the overexpression both of MR and Dectin-1. This upregulation requires arachidonic acid (AA) mobilization and the activation of peroxisome proliferator-activated receptor gamma (PPARγ) nuclear receptor through the leukotriene B4 (LTB4) production. AA/LTB4/PPARγ/Dectin-1-MR signaling pathway is crucial for P17-mediated anti-fungal activity of h-MDMs, as indicated by the fact that the activation of this axis by P17 triggered ROS production and inflammasome-dependent IL-1β release. Moreover, we showed that the increased anti-fungal immune response of h-MDMs by P17 was dependent on intracellular calcium mobilization triggered by the interaction of P17 with pertussis toxin-sensitive G-protein-coupled receptors on h-MDMs. Finally, we also demonstrated that P17-treated mice infected with C. albicans develop less severe gastrointestinal infection related to a higher efficiency of their macrophages to engulf Candida , to produce ROS and IL-1β and to kill the yeasts. Altogether, these results identify P17 as an original activator of the fungicidal response of macrophages that acts upstream PPARγ/CLRs axis and offer new immunomodulatory therapeutic perspectives in the field of infectious diseases.

Bees are exposed in their environment to contaminants that can weaken the colony and contribute to bee declines. Monoterpenoid-based preparations can be introduced into hives to control the parasitic mite Varroa destructor. The long-term effects of monoterpenoids are poorly investigated. Olfactory conditioning of the proboscis extension reflex (PER) has been used to evaluate the impact of stressors on cognitive functions of the honeybee such as learning and memory. The authors tested the PER to odorants on bees after exposure to monoterpenoids in hives. Octopamine receptors, transient receptor potential-like (TRPL), and γ-aminobutyric acid channels are thought to play a critical role in the memory of food experience. Gene expression levels of Amoa1, Rdl, and trpl were evaluated in parallel in the bee brain because these genes code for the cellular targets of monoterpenoids and some pesticides and neural circuits of memory require their expression. The miticide impaired the PER to odors in the 3 wk following treatment. Short-term and long-term olfactory memories were improved months after introduction of the monoterpenoids into the beehives. Chronic exposure to the miticide had significant effects on Amoa1, Rdl, and trpl gene expressions and modified seasonal changes in the expression of these genes in the brain. The decrease of expression of these genes in winter could partly explain the improvement of memory. The present study has led to new insights into alternative treatments, especially on their effects on memory and expression of selected genes involved in this cognitive function. Environ Toxicol Chem 2017;36:337-345. © 2016 SETAC., (© 2016 SETAC.)

We have recently characterized bicarinalin as the most abundant peptide from the venom of the ant Tetramorium bicarinatum. This antimicrobial peptide is active against Staphylococcus and Enterobacteriaceae. To further investigate the antimicrobial properties of this cationic and cysteine-free peptide, we have studied its antibacterial, antifungal and antiparasitic activities on a large array of microorganisms. Bicarinalin was active against fifteen microorganisms with minimal inhibitory concentrations ranging from 2 and 25μmolL(-1). Cronobacter sakazakii, Salmonella enterica, Candida albicans, Aspergilus niger and Saccharomyces cerevisiae were particularly susceptible to this novel antimicrobial peptide. Resistant strains of Staphylococcus aureus, Pseudomonas aeruginosa and C. albicans were as susceptible as the canonical strains. Interestingly, bicarinalin was also active against the parasite Leishmania infantum with a minimal inhibitory concentrations of 2μmolL(-1). The bicarinalin pre-propeptide cDNA sequence has been determined using a combination of degenerated primers with RACE PCR strategy. Interestingly, the N-terminal domain of bicarinalin pre-propeptide exhibited sequence similarity with the pilosulin antimicrobial peptide family previously described in the Myrmecia venoms. Moreover, using SYTOX green uptake assay, we showed that, for all the tested microorganisms, bicarinalin acted through a membrane permeabilization mechanism. Two dimensional-NMR experiments showed that bicarinalin displayed a 10 residue-long α-helical structure flanked by two N- and C-terminal disordered regions. This partially amphipathic helix may explain the membrane permeabilization mechanism of bicarinalin observed in this study. Finally, therapeutic value of bicarinalin was highlighted by its low cytotoxicity against human lymphocytes at bactericidal concentrations and its long half-life in human serum which was around 15h., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Essential oils are used by beekeepers to control the Varroa mites that infest honeybee colonies. So, bees can be exposed to thymol formulations in the hive. The effects of the monoterpenoid thymol were explored on olfactory memory and gene expression in the brain of the honeybee. In bees previously exposed to thymol (10 or 100 ng/bee), the specificity of the response to the conditioned stimulus (CS) was lost 24 h after learning. Besides, the octopamine receptor OA1 gene Amoa1 showed a significant decrease of expression 3 h after exposure with 10 or 100 ng/bee of thymol. With the same doses, expression of Rdl gene, coding for a GABA receptor subunit, was not significantly modified but the trpl gene was upregulated 1 and 24 h after exposure to thymol. These data indicated that the genes coding for the cellular targets of thymol could be rapidly regulated after exposure to this molecule. Memory and sensory processes should be investigated in bees after chronic exposure in the hive to thymol-based preparations.

Thymol is a natural substance increasingly used as an alternative to pesticides in the fight against the Varroa destructor mite. Despite the effectiveness of this phenolic monoterpene against Varroa, few articles have covered the negative or side effects of thymol on bees. In a previous study, we have found an impairment of phototaxis in honeybees following application of sublethal doses of thymol-lower or equal to 100 ng/bee-under laboratory conditions. The present work shows the same behavioral effects on bees from hives treated with Apilife Var®, a veterinary drug containing 74 % thymol, with a decrease in phototactic behavior observed 1 day after treatment. Thus, thymol causes disruption of bee phototactic behavior both under laboratory conditions as well as in beehives. The bee exposure dose in treated hives was quantified using gas chromatography coupled to mass spectrometry (GC-MS), giving a median value of 4.3 μg per body 24 h after treatment, with 11 ng in the brain. The thymol level in 20 organic waxes from hives treated with Apilife Var® was also measured and showed that it persists in waxes (around 10 mg/kg) 1 year after treatment. Thus, in the light of (1) behavioral data obtained under laboratory conditions and in beehives, (2) the persistence of thymol in waxes, and (3) the high load on bees, it would appear important to study the long-term effects of thymol in beehives.