Discovery of an Insect Neuroactive Helix Ring Peptide from Ant Venom.

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 U11, a 34-amino acid peptide, for further insecticidal, structural, and pharmacological experiments. Insecticidal assays revealed that U11 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 U11 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 U11 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.

Biological Activities and Ecological Significance of Fire Ant Venom Alkaloids.

Venoms produced by arthropods act as chemical weapons to paralyze prey or deter competitors. The utilization of venom is an essential feature in the biology and ecology of venomous arthropods. Solenopsis fire ants (Hymenoptera: Formicidae) are medically important venomous ants. They have acquired different patterns of venom use to maximize their competitive advantages rendered by the venom when facing different challenges. The major components of fire ant venom are piperidine alkaloids, which have strong insecticidal and antibiotic activities. The alkaloids protect fire ants from pathogens over the course of their lives and can be used to defend them from predators and competitors. They are also utilized by some of the fire ants' natural enemies, such as phorid flies to locate host ants. Collectively, these ants' diverse alkaloid compositions and functions have ecological significance for their survival, successful invasion, and rapid range expansion. The venom alkaloids with powerful biological activities may have played an important role in shaping the assembly of communities in both native and introduced ranges.

Synthesis and Insecticidal Activity of Fire Ant Venom Alkaloid-Based 2-Methyl-6-alkyl-Δ1,6-piperideines.

2,6-dialkylpiperideines found in the venom of Solenopsis (Hymenoptera, Formicidae) fire ants are a range of compounds possessing various biological activities. A series of racemic 2-methyl-6-alkyl-Δ1,6-piperideines were synthesized for chemical confirmation of the natural products found in fire ant venom, and the evaluation of their biological activity. Synthetic Δ1,6-piperideines and the natural compounds in the cis-alkaloid fraction of Solenopsis invicta had identical mass spectra and retention times. Their insecticidal activities against the third-instar larvae of cotton bollworm (Helicoverpa armigera) were evaluated by using injection and topical application methods. All three compounds exhibited no lethal effect at concentrations of 0.05-0.4 mol/L by topical treatment, but moderate lethal effect at 0.4 mol/L through injection treatment. Compound 6a showed significantly higher activity than the natural insecticide nicotine. The differences in activity among compounds 6b, 6c and nicotine were not significant. The elongation of the carbon chain at the 6-position of the piperideine ring appears to decrease insecticidal activity.

Multipurpose peptides: The venoms of Amazonian stinging ants contain anthelmintic ponericins with diverse predatory and defensive activities.

In the face of increasing drug resistance, the development of new anthelmintics is critical for controlling nematodes that parasitise livestock. Although hymenopteran venom toxins have attracted attention for applications in agriculture and medicine, few studies have explored their potential as anthelmintics. Here we assessed hymenopteran venoms as a possible source of new anthelmintic compounds by screening a panel of ten hymenopteran venoms against Haemonchus contortus, a major pathogenic nematode of ruminants. Using bioassay-guided fractionation coupled with liquid chromatography-tandem mass spectrometry, we identified four novel anthelmintic peptides (ponericins) from the venom of the neotropical ant Neoponera commutata and the previously described ponericin M-PONTX-Na1b from Neoponera apicalis venom. These peptides inhibit H. contortus development with IC50 values of 2.8-5.6 µM. Circular dichroism spectropolarimetry indicated that the ponericins are unstructured in aqueous solution but adopt α-helical conformations in lipid mimetic environments. We show that the ponericins induce non-specific membrane perturbation, which confers broad-spectrum antimicrobial, insecticidal, cytotoxic, hemolytic, and algogenic activities, with activity across all assays typically correlated. We also show for the first time that ponericins induce spontaneous pain behaviour when injected in mice. We propose that the broad-spectrum activity of the ponericins enables them to play both a predatory and defensive role in neoponeran ants, consistent with their high abundance in venom. This study reveals a broader functionality for ponericins than previously assumed, and highlights both the opportunities and challenges in pursuing ant venom peptides as potential therapeutics.

Dinoponera quadriceps venom as a source of active agents against Staphylococcus aureus.

Staphylococcus aureus is a highly virulent pathogen, capable of biofilm formation and responsible for thousands of deaths each year. The prevalence of Methicillin-Resistant S. aureus (MRSA) strains has increased in recent years and thus, the development of new antibiotics has become necessary. Antimicrobial Peptides (AMPs) are effective against a variety of multidrug-resistant bacteria and low levels of resistance have been reported regarding these molecules. Dinoponera quadriceps ant venom (DqV) has been described regarding its effect against S. aureus. In this study, we have evaluated the antibacterial effect of DqV-AMPs, the dinoponeratoxins (DNTxs), against Methicillin-Sensitive and a Methicillin-Resistant S. aureus strains. Our results show DNTx M-PONTX-Dq3a as a potent inhibitor of both strains, being able to prevent biofilm formation at low micromolar range (0.78-3.12 µM). It also showed a short-time effect through membrane disruption. M-PONTX-Dq3a opens up new perspectives for the prevention of biofilm formation through the development of anti-adhesive surface coatings on medical devices, as well as the treatment of resistant strains in skin or soft tissue infections.

Identification and Functional Characterization of a Novel Insecticidal Decapeptide from the Myrmicine Ant Manica rubida.

Ant venoms contain many small, linear peptides, an untapped source of bioactive peptide toxins. The control of agricultural insect pests currently depends primarily on chemical insecticides, but their intensive use damages the environment and human health, and encourages the emergence of resistant pest populations. This has promoted interest in animal venoms as a source of alternative, environmentally-friendly bio-insecticides. We tested the crude venom of the predatory ant, Manica rubida, and observed severe fitness costs in the parthenogenetic pea aphid (Acyrthosiphon pisum), a common agricultural pest. Therefore, we explored the M. rubida venom peptidome and identified a novel decapeptide U-MYRTX-MANr1 (NH2-IDPKVLESLV-CONH2) using a combination of Edman degradation and de novo peptide sequencing. Although this myrmicitoxin was inactive against bacteria and fungi, it reduced aphid survival and reproduction. Furthermore, both crude venom and U-MYRTX-MANr1 reversibly paralyzed injected aphids and induced a loss of body fluids. Components of M. rubida venom may act on various biological targets including ion channels and hemolymph coagulation proteins, as previously shown for other ant venom toxins. The remarkable insecticidal activity of M. rubida venom suggests it may be a promising source of additional bio-insecticide leads.

Fire Ant Venom Alkaloids Inhibit Biofilm Formation.

Biofilm formation on exposed surfaces is a serious issue for the food industry and medical health facilities. There are many proposed strategies to delay, reduce, or even eliminate biofilm formation on surfaces. The present study focuses on the applicability of fire ant venom alkaloids (aka 'solenopsins', from Solenopsis invicta) tested on polystyrene and stainless steel surfaces relative to the adhesion and biofilm-formation by the bacterium Pseudomonas fluorescens. Conditioning with solenopsins demonstrates significant reduction of bacterial adhesion. Inhibition rates were 62.7% on polystyrene and 59.0% on stainless steel surfaces. In addition, solenopsins drastically reduced cell populations already growing on conditioned surfaces. Contrary to assumptions by previous authors, solenopsins tested negative for amphipathic properties, thus understanding the mechanisms behind the observed effects still relies on further investigation.

A comprehensive portrait of the venom of the giant red bull ant, Myrmecia gulosa, reveals a hyperdiverse hymenopteran toxin gene family.

Ants (Hymenoptera: Formicidae) are diverse and ubiquitous, and their ability to sting is familiar to many of us. However, their venoms remain largely unstudied. We provide the first comprehensive characterization of a polypeptidic ant venom, that of the giant red bull ant, Myrmecia gulosa. We reveal a suite of novel peptides with a range of posttranslational modifications, including disulfide bond formation, dimerization, and glycosylation. One venom peptide has sequence features consistent with an epidermal growth factor fold, while the remaining peptides have features suggestive of a capacity to form amphipathic helices. We show that these peptides are derived from what appears to be a single, pharmacologically diverse, gene superfamily (aculeatoxins) that includes most venom peptides previously reported from the aculeate Hymenoptera. Two aculeatoxins purified from the venom were found to be capable of activating mammalian sensory neurons, consistent with the capacity to produce pain but via distinct mechanisms of action. Further investigation of the major venom peptide MIITX1-Mg1a revealed that it can also incapacitate arthropods, indicative of dual utility in both defense and predation. MIITX1-Mg1a accomplishes these functions by generating a leak in membrane ion conductance, which alters membrane potential and triggers neuronal depolarization. Our results provide the first insights into the evolution of the major toxin gene superfamily of the aculeate Hymenoptera and provide a new paradigm in the functional evolution of toxins from animal venoms.

Mechanistic and phenotypic studies of bicarinalin, BP100 and colistin action on Acinetobacter baumannii.

Acinetobacter baumannii has been identified by the WHO as a high priority pathogen. It can be resistant to multiple antibiotics and colistin sulphate is often used as a last-resort treatment. However, the potentially severe side-effects of colistin are well documented and this study compared the bactericidal and anti-biofilm activity of two synthetic nature-inspired antimicrobial peptides, bicarinalin and BP100, with colistin. The minimum bactericidal concentration (MBC) against planktonic A. baumannii was approximately 0.5 µg/ml for colistin sulphate and ∼4 µg/ml for bicarinalin and BP100. A. baumannii commonly occurs as a biofilm and biofilm removal assay results highlighted that both bicarinalin and BP100 had significantly greater potential than colistin. Atomic force microscopy (AFM) showed dramatic changes in A. baumannii cell size and surface conformity when treated with peptide concentrations at and above the MBC. Scanning electron microscopy (SEM) visualised the reduction of biofilm coverage and cell surface changes as peptide concentration increased. Liposome assays revealed that these peptides most likely act as pore-forming agents in the membrane. Bicarinalin and BP100 may be effective therapeutic alternatives to colistin against A. baumannii infections but further research is required to assess if they elicit cytotoxicity issues in patients.

Samsum Ant Venom Exerts Anticancer Activity Through Immunomodulation In Vitro and In Vivo.

Samsum ant venom (SAV) is a rich repertoire of natural compounds with tremendous pharmacological properties. The present work explores its antineoplastic activity in different cell lines followed by its confirmation in vivo. The cell lines, HepG2, MCF-7, and LoVo showed the differential dose-dependent antineoplastic effect with an increased level of significant cytokines, including Interleukin (IL)-1ß, IL-6, and IL-8 and transcription factor, Nuclear factor-kappa B (NF-κB). However, the venom was more effective on HepG2 and MCF-7 cells than LoVo cells. Furthermore, the extract was administered to four groups (n = 8) of rats. Group I was taken as a control without any treatment, whereas group II received CCl4 (1 mL/kg) for induction of mild hepatoma. Group III was given 100 µg/kg of SAV twice a week for 1 month. Group IV was pretreated with the CCl4 (like group II) followed by dosing with SAV (100 µg/kg) for 2 months as per the authors' prestandardized dosing schedule. Intriguingly, the rats of group IV demonstrated significant decrease in key cytokines, IL-1ß and IL-6, as well as the transcription factors, including Tumor Necrosis Factor-alpha (TNF-α), NF-κB, and Inhibitor-kappa B (I-κB) as compared with group II. Furthermore, increase in IL-10 and First apoptosis signal (FAS) in the same group confirmed that SAV induces apoptosis at the given dose through immunomodulation leading to enhanced tumor killing in vivo. Hence, SAV has an excellent antineoplastic activity that can be directly used to treat certain types of cancer. Moreover, study of its ingredients can pave ways to design novel anticancer drugs. However, further in-depth investigation is required before its clinical trials.

Anti-Helicobacter pylori Properties of the Ant-Venom Peptide Bicarinalin.

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 IC50 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.

Synthesis and Antimicrobial Evaluation of Fire Ant Venom Alkaloid Based 2-Methyl-6-alkyl-Δ1,6-piperideines.

The first synthesis of 2-methyl-6-pentadecyl-Δ1,6-piperideine (1), a major alkaloid of the piperideine chemotype in fire ant venoms, and its analogues, 2-methyl-6-tetradecyl-Δ1,6-piperideine (2) and 2-methyl-6-hexadecyl-Δ1,6-piperideine (3), was achieved by a facile synthetic method starting with glutaric acid (4) and urea (5). Compound 1 showed in vitro antifungal activity against Cryptococcus neoformans and Candida albicans with IC50 values of 6.6 and 12.4 µg/mL, respectively, and antibacterial activity against vancomycin-resistant Enterococcus faecium with an IC50 value of 19.4 µg/mL, while compounds 2 and 3 were less active against these pathogens. All three compounds strongly inhibited the parasites Leishmania donovani promastigotes and Trypanosoma brucei with IC50 values in the range of 5.0-6.7 and 2.7-4.0 µg/mL, respectively.

A reexamination of poneratoxin from the venom of the bullet ant Paraponera clavata.

In 1991, Piek et al. [45] described a voltage-gated sodium channel (VGSC) modifier from "bullet ant" (Paraponera clavata) venom they called poneratoxin (PoTx). Using UV chromatography and Edman degradation they showed two "identical peptides" of 25 residues. We reinvestigated PoTx using ultra performance liquid chromatography-tandem mass spectrometry (UPLC-TMS). De novo sequencing showed the two peptides were actually structurally different peptides: the originally described PoTx and a glycyl pro-peptide (glycyl-PoTx) that lacks C-terminus amidation. We examined P. clavata venom from different geographical locations and discovered two additional PoTx analogs: an A23E substitution analog and a D22N; A23V substitutions analog. We tested PoTx and these three natural analogs on the mammalian sensory voltage-gated sodium channel, Nav1.7, using whole cell voltage-clamp. PoTx and each analog induced slowly activating currents in response to small depolarizing steps and sustained currents due to blockade of channel inactivation, similar to that described previously in skeletal muscle [19]. Glycyl-PoTx had the same potency and efficacy as PoTx. A23E PoTx, with a decrease in both C-terminal net positive charge and hydrophobicity, had an eight-fold reduction in potency compared to PoTx. In contrast, the D22N; A23V PoTx, with an increase in both C-terminal net positive charge and hydrophobicity, had a nearly five-fold increase in potency compared to PoTx. We found that changes in PoTx C-terminus caused a significant change in PoTx potency.

Anticonvulsant Effects of Fractions Isolated from Dinoponera quadriceps (Kempt) Ant Venom (Formicidae: Ponerinae).

Natural products, sources of new pharmacological substances, have large chemical diversity and architectural complexity. In this context, some toxins obtained from invertebrate venoms have anticonvulsant effects. Epilepsy is a neurological disorder that affects about 65 million people worldwide, and approximately 30% of cases are resistant to pharmacological treatment. Previous studies from our group show that the denatured venom of the ant Dinoponera quadriceps (Kempt) protects mice against bicuculline (BIC)-induced seizures and death. The aim of this study was to investigate the anticonvulsant activity of compounds isolated from D. quadriceps venom against seizures induced by BIC in mice. Crude venom was fractionated by high-performance liquid chromatography (HPLC) resulting in six fractions referred to as DqTx1-DqTx6. A liquid chromatography-mass spectrometry (LC/MS) analysis revealed a major 431 Da compound in fractions DqTx1 and DqTx2. Fractions DqTx3 and DqTx4 showed a compound of 2451 Da and DqTx5 revealed a 2436 Da compound. Furthermore, the DqTx6 fraction exhibited a major component with a molecular weight of 13,196 Da. Each fraction (1 mg/mL) was microinjected into the lateral ventricle of mice, and the animals were observed in an open field. We did not observe behavioral alterations when the fractions were given alone. Conversely, when the fractions were microinjected 20 min prior to the administration of BIC (21.6 nM), DqTx1, DqTx4, and DqTx6 fractions increased the latency for onset of tonic-clonic seizures. Moreover, all fractions, except DqTx5, increased latency to death. The more relevant result was obtained with the DqTx6 fraction, which protected 62.5% of the animals against tonic-clonic seizures. Furthermore, this fraction protected 100% of the animals from seizure episodes followed by death. Taken together, these findings indicate that compounds from ant venom might be a potential source of new anticonvulsants molecules.

Potential effects of samsum ant, Brachyponera sennaarensis, venom on TNF-α/NF-κB mediated inflammation in CCL4-toxicity in vivo.

BACKGROUND: Ant venom shows antimicrobial, anti-parasitic and anti-inflammatory activities, both in vitro and in vivo. Our recent studies have confirmed the role of samsum ant venom (SAV) as a powerful antioxidant. This study aimed to investigate whether SAV as a potential treatment for CCl4-induced acute liver toxicity in an animal (rat) model. METHODS: Thirty-two rats were assigned into four groups; the first one served as the control. The second group received a single dose of 1 ml/kg CCl4 in a 1:1 ratio with olive oil through an intraperitoneal injection. The third group received a single dose of 1 ml/kg CCl4 and then treated with SAV at a dose of 100 µg SAV twice a week for three weeks. The fourth group received a dose of 100 µg SAV only twice a week for three weeks. ELISA, RT-PCR and histopathological examinations were applied. RESULTS: Results showed that antioxidant enzymes were significantly reduced in the diseased animals. SAV was found to significantly restore the oxidative stability in diseased animals. ELISA estimation and RT-PCR analysis also showed significant upregulation of both nuclear factor (κB) NF-κB and inhibitor (κB) IκB, respectively, in the diseased animals compared to the normal ones. The expression of tumour necrosis factor alpha (TNF-α) and pro-apoptotic receptor (Fas) were also significantly up-regulated in the diseased rats. Interestingly, SAV was found to significantly restore NF-κB, IκB and TNF-α in the diseased rats to the normal values. As a result, liver enzymes, serum proteins and lipid concentrations were significantly improved by SAV in CCl4-animals in comparison with the control ones. Moreover, SAV obviously improved the hepatic tissues of the same group was. CONCLUSION: SAV treatment restores the normal biochemical and oxidative stability by improving the TNF-α/NF-κB mediated inflammation in CCL4-treated rats.

Biochemical and biophysical combined study of bicarinalin, an ant venom antimicrobial peptide.

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.

Analysis of Protein Composition and Bioactivity of Neoponera villosa Venom (Hymenoptera: Formicidae).

Ants cause a series of accidents involving humans. Such accidents generate different reactions in the body, ranging from a mild irritation at the bite site to anaphylactic shock, and these reactions depend on the mechanism of action of the venom. The study of animal venom is a science known as venomics. Through venomics, the composition of the venom of several ant species has already been characterized and their biological activities described. Thus, the aim of this study was to evaluate the protein composition and biological activities (hemolytic and immunostimulatory) of the venom of Neoponera villosa (N. villosa), an ant widely distributed in South America. The protein composition was evaluated by proteomic techniques, such as two-dimensional electrophoresis. To assess the biological activity, hemolysis assay was carried out and cytokines were quantified after exposure of macrophages to the venom. The venom of N. villosa has a profile composed of 145 proteins, including structural and metabolic components (e.g., tubulin and ATPase), allergenic and immunomodulatory proteins (arginine kinase and heat shock proteins (HSPs)), protective proteins of venom (superoxide dismutase (SOD) and catalase) and tissue degradation proteins (hyaluronidase and phospholipase A2). The venom was able to induce hemolysis in human erythrocytes and also induced release of both pro-inflammatory cytokines, as the anti-inflammatory cytokine release by murine macrophages. These results allow better understanding of the composition and complexity of N. villosa venom in the human body, as well as the possible mechanisms of action after the bite.

Novel Pharmacological Properties of Dinoponera quadriceps Giant Ant Venom.

The South American giant ant, Dinoponera quadriceps (Hymenoptera, Formicidae, Ponerinae), produces proteinaceous venom that has antinociceptive, neuroprotective and antimicrobial effects, thereby supporting the popular use of these ants to treat asthma, rheumatism, earache and back pain. Anticoagulant activity is another biological property that has been shown for the venom of other hymenopteran species, like wasps. The aim of this study was to assess the anti-inflammatory, anticoagulant and antiplatelet properties of D. quadriceps venom (DqV). DqV anti-inflammatory activity was assessed by intravenous administration in Swiss mice in the models of paw edema and peritonitis. In vitro, DqV was assessed in coagulation (activated partial thromboplastin time) and platelet aggregation tests. DqV inhibited (27-33%) the edema elicited by carrageenan and the leucocyte migration (43%) elicited by zymosan. DqV decreased by 57% and 42%, respectively, the content of malondialdehyde and nitrite in the peritoneal fluid. DqV prolonged (1.8x) the clotting time and decreased (27%) the platelet aggregation induced by adenosine diphosphate. The crude venom of D. quadriceps presents an anti-inflammatory effect in mice and in vitro anticoagulant and antiplatelet effects.

Linear antimicrobial peptides from Ectatomma quadridens ant venom.

Venoms from three poneromorph ant species (Paraponera clavata, Ectatomma quadridens and Ectatomma tuberculatum) were investigated for the growth inhibition of Gram-positive and Gram-negative bacteria. It was shown that the venom of E. quadridens and its peptide fraction in particular possess marked antibacterial action. Three linear antimicrobial peptides sharing low similarity to the well-known ponericin peptides were isolated from this ant venom by means of size-exclusion and reversed-phase chromatography. The peptides showed antimicrobial activity at low micromolar concentrations. Their primary structure was established by direct Edman sequencing in combination with mass spectrometry. The most active peptide designated ponericin-Q42 was chemically synthesized. Its secondary structure was investigated in aqueous and membrane-mimicking environment, and the peptide was shown to be partially helical already in water, which is unusual for short linear peptides. Analysis of its activity on different bacterial strains, human erythrocytes and chronic myelogenous leukemia K562 cells revealed that the peptide shows broad spectrum cytolytic activity at micromolar and submicromolar concentrations. Ponericin-Q42 also possesses weak toxic activity on flesh fly larvae with LD50 of ∼105 µg/g.

Antimicrobial effect of Dinoponera quadriceps (Hymenoptera: Formicidae) venom against Staphylococcus aureus strains.

AIMS: Dinoponera quadriceps venom (DqV) was examined to evaluate the antibacterial activity and its bactericidal action mechanism against Staphylococcus aureus. METHODS AND RESULTS: DqV was tested against a standard strain of methicillin-sensitive Staphylococcus aureus (MSSA), Staph. aureus ATCC 6538P and two standard strains of methicillin-resistant Staphylococcus aureus (MRSA), Staph. aureus ATCC 33591 and Staph. aureus CCBH 5330. The minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC), the rate of kill and pH sensitivity of the DqV were determined by microdilution tests. Bactericidal and inhibitory concentrations of DqV were tested to check its action on Staph. aureus membrane permeability and cell morphology. The MIC and MBC of DqV were 6·25 and 12·5 µg ml(-1) for Staph. aureus ATCC 6538P, 12·5 and 50 µg ml(-1) for Staph. aureus CCBH 5330 and 100 and 100 µg ml(-1) for Staph. aureus ATCC 33591, respectively. Complete bacterial growth inhibition was observed after 4 h of incubation with the MBC of DqV. A lowest MIC was observed in alkaline pH. Alteration in membrane permeability was observed through the increase in crystal violet uptake, genetic material release and morphology in atomic force microscopy. CONCLUSIONS: The results suggest antibacterial activity of DqV against Staph. aureus and that the venom acts in the cell membrane. SIGNIFICANCE AND IMPACT OF THE STUDY: Alteration in membrane permeability may be associated with the antimicrobial activity of hymenopteran venoms.