melitten and Insect-Bites-and-Stings

Episodic hemorrhage is not a typical symptom of anaphylactic reaction to insect stings. Cases of reactions to honeybee (HB) sting or venom immunotherapy in which the uterus is the main target organ are very rare. Hemorrhage can be induced by HB venom components, especially melittin, which interfere with complement cleavage and bradykinin release. Both mechanisms are directly or indirectly associated with coagulation, thrombolysis, hemolysis, and smooth muscle tone. Induction of episodic hemorrhage through pathway destabilization in a defective bradykinin system or vulnerable organ may not be compensated by appropriate regulatory mechanisms. The pathological role of effectors is generally offset by the interaction of various regulatory systems, and the probability of hemorrhage is minimized thanks to this compensatory capability. In endometrial bleeding, the uterus becomes more vulnerable as a result of postmenstrual vascular fragility and additional induction of anaphylaxis-related uterine contractions. Episodic hemorrhage, especially metrorrhagia, as a consequence of HB venom activity may be suspected by an allergologist, but not by a physician. Melittin-free or recombinant allergens of HB venom, as well as modulators of the biochemical systems involved, could help to reduce the likelihood of hemorrhage. However, further investigation is required before these strategies can be introduced in clinical practice.

Topics: Anaphylaxis; Animals; Bee Venoms; Bees; Bites and Stings; Bradykinin; Complement System Proteins; Female; Humans; Insect Bites and Stings; Melitten; Metrorrhagia; Uterus

Bee stings represent a public health subject, but the mechanisms involved in bee venom toxicity are not yet fully understood. To evaluate the reactions of adrenocortical cells, through which organisms respond to stress, two honeybee venom components: melittin (Mlt) and phospholipase A2 (PLA2) were tested as potential chemical stressors. Modifications were investigated with transmission electron microscopy and microanalysis. A single dose of Mlt (31 mg/kg) or PLA2 (9.3 mg/kg) was injected in rats of groups ML and PL; daily doses of Mlt (350 μg/kg) or PLA2 (105 μg/kg) were injected 30 days in rats of groups M30 and P30. Adrenocortical cells in ML group showed ultrastructural degenerative alterations of nuclei, endoplasmic reticulum, and mitochondria that exhibited lipid inclusions and mitochondrial cristae (MC) re-organized into mono- or multimembrane large vesicles, and whorls of membranes. Many MC were degenerated. In the M30 group, similar ultrastructural changes, but of lower amplitude were noted; lipid cytosolic droplets were heterogenous. MC diameters in Mlt groups (melittin treated groups) were significantly higher than in control (C) group. In PL group, mitochondria contained large lipid inclusions, vesicular MC of different sizes and multiple membranes, and debris, or whorl structures. In P30 group MC were tubular with increased diameters. In both PLA2 groups (PLA2 treated groups) MC were significantly larger than in C group. We concluded that Mlt and PLA2 were powerful stressors, toxic at the tested doses, cellular reactions concerning in all groups mainly mitochondria, but also other cellular compartments. Apart from degenerative regression of MC, the rearrangement of tubular MC occurred into one or multiple large multimembrane vesicular MC. Reactions to the high doses were more pronounced, with the highest amplitude in ML group, and the lowest in P30 group.

Topics: Animals; Bee Venoms; Bees; Insect Bites and Stings; Lipids; Melitten; Mitochondria; Phospholipases A2; Rats

Topics: Animals; Antibodies, Neutralizing; Bee Venoms; Bees; Chick Embryo; Chickens; Egg Yolk; Female; Immunoglobulins; Insect Bites and Stings; Male; Melitten; Mice; Phospholipases A2

A bee sting can result in allergic and toxin-mediated local manifestations like pain, swelling, redness and itching to serious systemic effects like acute kidney injury (AKI), pancreatitis, Kounis syndrome and stroke. Melittin and phospholipase A2, which make up 62% of honeybee venom, have vasoactive, haemolytic properties causing severe AKI. Its role in lowering blood glucose in diabetics is an interesting research topic. We report an elderly herdsman, a known diabetic on irregular oral hypoglycaemic drugs, who presented with altered mental status due to hypoglycaemia. On further prodding, a recent multiple bee sting attack 5 days ago was found which was followed by altered coloured urine for 2 days for which no medical attention was sought. Additional analyses revealed reticulocytosis, azotemia and high serum creatine phosphokinase. The patient was treated with dextrose infusion, antihistamines, fluids and haemodialysis. Renal failure resolved completely and the patient was discharged in a stable condition.

Topics: Aged; Allergens; Animals; Bee Venoms; Bees; Humans; Hypoglycemia; Insect Bites and Stings; Melitten

Bees defend themselves by stinging and injecting a venom into their victims; bee venom is a complex mixture of chemicals including the polypeptide melittin which is mainly responsible for triggering the pain of the sting.

Topics: Animals; Bee Venoms; Bees; Insect Bites and Stings; Melitten

Africanized Apis mellifera bees, also known as killer bees, have an exceptional defensive instinct, characterized by mass attacks that may cause envenomation or death. From the years 2000-2013, 77,066 bee accidents occurred in Brazil. Bee venom comprises several substances, including melittin and phospholipase A2 (PLA2). Due to the lack of antivenom for bee envenomation, this study aimed to produce human monoclonal antibody fragments (single chain fragment variable; scFv), by using phage display technology. These fragments targeted melittin and PLA2, the two major components of bee venom, to minimize their toxic effects in cases of mass envenomation. Two phage antibody selections were performed using purified melittin. As the commercial melittin is contaminated with PLA2, phages specific to PLA2 were also obtained during one of the selections. Specific clones for melittin and PLA2 were selected for the production of soluble scFvs, named here Afribumabs: prefix: afrib- (from Africanized bee); stem/suffix: -umab (fully human antibody). Afribumabs 1 and 2 were tested in in vitro and in vivo assays to assess their ability to inhibit the toxic actions of purified melittin, PLA2, and crude bee venom. Afribumabs reduced hemolysis caused by purified melittin and PLA2 and by crude venom in vitro and reduced edema formation in the paws of mice and prolonged the survival of venom-injected animals in vivo. These results demonstrate that Afribumabs may contribute to the production of the first non-heterologous antivenom treatment against bee envenomation. Such a treatment may overcome some of the difficulties associated with conventional immunotherapy techniques.

Topics: Animals; Antivenins; Bee Venoms; Cell Surface Display Techniques; Clone Cells; Drug Design; Drug Therapy, Combination; Edema; Hemolysis; Humans; Insect Bites and Stings; Insect Proteins; Male; Melitten; Mice; Phospholipase A2 Inhibitors; Phospholipases A2, Secretory; Recombinant Proteins; Single-Chain Antibodies; Subcutaneous Tissue; Survival Analysis

The hybrid created from the crossbreeding of European and African bees, known as the Africanised bee, has provided numerous advantages for current beekeeping. However, this new species exhibits undesirable behaviours, such as colony defence instinct and a propensity to attack en masse, which can result in serious accidents. To date, there is no effective treatment for cases of Africanised bee envenomation. One promising technique for developing an efficient antivenom is the use of phage display technology, which enables the production of human antibodies, thus avoiding the complications of serum therapy, such as anaphylaxis and serum sickness. The aim of this study was to produce human monoclonal single-chain Fv (scFv) antibody fragments capable of inhibiting the toxic effects of Africanised bee venom. We conducted four rounds of selection of antibodies against the venom and three rounds of selection of antibodies against purified melittin. Three clones were selected and tested by enzyme-linked immunosorbent assay to verify their specificity for melittin and phospholipase A2. Two clones (C5 and C12) were specific for melittin, and one (A7) was specific for phospholipase A2. In a kinetic haemolytic assay, these clones were evaluated individually and in pairs. The A7-C12 combination had the best synergistic effect and was chosen to be used in the assays of myotoxicity inhibition and lethality. The A7-C12 combination inhibited the in vivo myotoxic effect of the venom and increased the survival of treated animals.

Topics: Animals; Antibodies, Monoclonal; Antivenins; Bee Venoms; Bees; Enzyme-Linked Immunosorbent Assay; Female; Humans; Insect Bites and Stings; Melitten; Mice; Phospholipases A2; Single-Chain Antibodies; Survival

Bee stings are a health concern in the Americas, where fatal envenomings due to massive attacks by Africanized honeybees have been documented in the last decades. Most studies on the toxic effects of honeybee venom in experimental animals have been performed using the intravenous or intraperitoneal injection routes. The aim of this study was to develop a mouse model that would better resemble a massive honeybee attack by using the subcutaneous (s.c.) route to induce a severe, sublethal systemic envenoming. An array of acute venom effects were characterized, including biochemical, hematological, histological, and inflammatory alterations, after the s.c. injection of 0.5 median lethal dose of venom. Rapid increases in serum alanine (ALT) and aspartate (AST) transaminases, creatinine, urea nitrogen, uric acid, sodium and chloride electrolytes, and creatine kinase (CK) were recorded, indicating damage to liver, kidneys, and skeletal muscle. Also, coagulation disturbances (fibrinogen decrease, and moderate delay in prothrombin and partial thromboplastin times) were demonstrated. Circulating platelet and leukocyte numbers remained unaltered, but a hemoconcentration effect (hematocrit and hemoglobin increase) was observed. This effect might be related to the marked edema induced by the venom. In addition, this inflammatory response included a systemic increase in cytokines (IL-1 beta, IL-6, TNF-alpha), together with an elevation of serum malondialdehyde and nitric oxide. The myotoxic effects of venom, melittin, and phospholipase A(2) were demonstrated after injection by s.c. route. No synergistic myotoxicity between melittin and PLA(2) was observed. Moreover, these two components, when injected at equivalent concentrations to those present in venom, induced a lower increase in serum CK than venom, suggesting that other components also contribute to its strong systemic toxicity towards skeletal muscle. The model here presented may be useful in preclinical studies to assess therapeutic antivenoms developed to cope with the problem of massive bee attacks.

Topics: Alanine Transaminase; Animals; Aspartate Aminotransferases; Bee Venoms; Bees; Biomarkers; Blood Coagulation; Cytokines; Disease Models, Animal; Drug Synergism; Female; Injections, Subcutaneous; Insect Bites and Stings; Lethal Dose 50; Male; Melitten; Mice; Mice, Inbred Strains; Myositis; Oxidative Stress; Phospholipases A2

The lethal effects of Africanized honey bee venom depend on the absorption of venom delivered during simultaneous sting attacks by large numbers of bees. The hypothesis that antibodies to whole bee venom and bee venom components could neutralize the lethal effect of bee venom was tested. Antibodies from beekeepers and immunized rabbits were incubated with bee venom and neutralization was studied by survival of intravenously injected mice. Beekeeper serum antibodies were found effective in protecting mice challenged with whole venom, and serum from rabbits immunized with phospholipase A2 (PLA2) was effective in protection against lethal effects of PLA2. Serum antibodies from rabbits immunized with whole venom or melittin were ineffective in neutralizing whole venom in vivo and had low titers in a venom enzyme-linked immunosorbent assay. The results suggest the need for development of more effective methods for raising antitoxic antibodies to bee venom components in other animals as a means of developing an antiserum that would be effective for treatment of human victims of multiple bee stings.

Topics: Animals; Antivenins; Bee Venoms; Bees; Dose-Response Relationship, Immunologic; Humans; Immune Sera; Immunoglobulin G; Insect Bites and Stings; Lethal Dose 50; Melitten; Mice; Phospholipases A; Phospholipases A2; Rabbits

The venoms of Apis dorsata, A. cerana, A. florea, and three different populations of A. mellifera were compared for lethal activity toward mice. All venoms exhibited identical activities, a finding consistent with recent evolutionary history within the genus. Young queen honeybees use their venoms only for stinging other queens and possess a venom only half as lethal to mice as worker venom, and by the time queens are 1-2 years of age their venom has become essentially inactive. Phospholipase A2 is the most lethal of the honeybee venom peptides, whereas melittin, which is only slightly less lethal, is the most abundant. Concurrent analyses of melittin, phospholipase, and the combination of the two at their natural 3:1 mixture in bee venom revealed that the lethal activity of the mixture was about the same as native honeybee venom. This value was less than that for either melittin or phospholipase alone and indicates that synergism of the two peptides is not occurring. The results are consistent with independent lethal activities for the venom components, and show that melittin is not only the dominant, but also the main lethal component in honeybee venom.

Topics: Animals; Apamin; Bee Venoms; Insect Bites and Stings; Melitten; Mice; Phospholipases A; Phospholipases A2; Species Specificity; Toxins, Biological

To determine the rate and completeness of delivery of venom from honeybee stings, European bees were collected at the entrance of a hive and studied with the use of two laboratory models. In one model bees were induced to sting the shaved skin of anesthetized rabbits. The stings were removed from the skin at various time intervals after autotomization, and residual venom was assayed with a hemolytic method. In the other model the bees were induced to sting preweighed filter paper disks, which were weighed again after removal of the sting at various intervals. Results of both experiments were in agreement, showing that at least 90% of the venom sac contents were delivered within 20 seconds and that venom delivery was complete within 1 minute. The data suggest that a bee sting must be removed within a few seconds after autotomization to prevent anaphylaxis in an allergic person. The extensive variation found in the amount of venom delivered at each time point may explain inconsistencies in relationships among reactions to field stings, sting challenge testing, venom skin tests and RAST.

Topics: Anaphylaxis; Animals; Bee Venoms; Bees; Insect Bites and Stings; Melitten; Rabbits; Skin; Time Factors

IgE antibodies to purified proteins and peptides from honeybee venom have been measured by the RAST. Trace amounts (less than 0.1%) of the major venom protein phospholipase A2 (PLA2) grossly distorted the measurement of IgE antibody to the other venom proteins, acid phosphatase (Acid P) and hyaluronidase (HYAL), and overemphasized their importance. Reduction of antigen coupled to the cellulose paper discs, which were used in the assay, diluted out the contaminating PLA2 without apparent loss in sensitivity. The reduction of disc-bound antigen increased the competition between IgE and IgG antibodies but did not affect measurement of IgE antibodies in sera taken from 35 untreated patients who had a history of general allergic reactions to bee stings. In 54% of sera from bee venom--allergic patients, the greatest IgE antibody response was to PLA2. In all, IgE antibodies to PLA2 were present in 91% of these sera. IgE antibodies to Acid P, HYAL, or melittin were present in 60%, 51%, and 31% of sera, respectively, and accounted for the highest level of binding in 17%, 17%, and 6% of these. Only 6% of sera were positive for whole venom but negative for the isolated antigens. A low level of IgE antibody was found to peptide 401 in 6% of sera. No IgE antibodies were found to apamin. While confirming the central role played by PLA2 in bee sting allergy, these results show that other venom components are also important in some patients.

Topics: Acid Phosphatase; Antibodies; Antibodies, Anti-Idiotypic; Antibody Specificity; Apamin; Bee Venoms; Bees; Humans; Hyaluronoglucosaminidase; Hypersensitivity; Immunoglobulin E; Immunoglobulin G; Insect Bites and Stings; Melitten; Phospholipases A; Phospholipases A2; Radioallergosorbent Test

Topics: Animals; Ant Venoms; Apamin; Bee Venoms; Female; Humans; Hymenoptera; Hypersensitivity; Insect Bites and Stings; Melitten; Venoms; Wasps

A group of 25 honey bee venom allergic patients were treated with commercial honey bee venom at a monthly maintenance dose of 100 micrograms for approximately one year. At the end of one year 24 patients were intentionally challenged and one was accidentally challenged. Three patients experienced significant systemic reactions to challenge and three experienced minor reactions. Sera obtained before commencing therapy, at maintenance and before challenge were tested by radioallergosorbent test (RAST), double antibody technique and protein A RAST for IgE and IgG antibody levels to all five known honey bee venom allergens. All of the treatment failures experienced at least a two-fold rise in IgG antibody against phospholipase. The ratios of IgG to IgE antibodies in the pre-challenge specimens were analyzed by a graphical method. Four patients had inadequate responses to at least three of the five allergens and three of these patients were those who experienced severe reactions to challenge. Sixteen patients had adequate responses to all five allergens, four patients to four allergens and one patient to three allergens; three of these patients experienced minor or local reactions to challenge and the remainder no reaction. No single allergen identified only the three severe reactors but three allergens identified all three reactors. The diagnostic efficiency of the criteria used for assessing protection was 0.96. The only non-correlating case was classifying a single nonreactor as at risk. No patients were misclassified as protected.

Topics: Allergens; Bee Venoms; Humans; Hyaluronoglucosaminidase; Immunoglobulin E; Immunoglobulin G; Insect Bites and Stings; Melitten; Phospholipases A; Phosphoric Monoester Hydrolases; Radioallergosorbent Test

Specific IgE antibodies against bee venom and its components were studied in 23 bee-keepers. The highest IgG serum levels were observed for whole bee venom followed by phospholipase A. The serum levels of specific IgG antibodies against melittin and MCD-peptide were lower, the lowest serum levels being observed for apamin. After a 5 month absence from bee-keeping a fall in the serum levels of IgG antibodies was observed in all the bee-keepers studied. The investigation of the IgG subclass antibodies 1-4 against bee venom and phospholipase A demonstrated the highest serum levels for IgG 4 and IgG 2, the lowest levels were observed for IgG 1. The lowest IgG serum levels were associated with the least effective protection to bee stings. These findings support the concept that specific IgG antibodies prevent the development of allergic symptoms after bee sting.

Topics: Adult; Aged; Apamin; Bee Venoms; Bees; Humans; Immunoglobulin G; Insect Bites and Stings; Melitten; Middle Aged; Peptides; Phospholipases A

Specific IgE antibodies against bee venom, phospholipase A, melittin and wasp venom have been examined in fifty patients with an unusually severe reaction after bee or wasp sting. Two thirds of the bee venom-sensitive patients also have detectable IgE antibodies to wasp venom. More than 50% of the wasp venom-sensitive patients are also allergic to bee venom. Phospholipase A and melittin IgE antibodies were found, respectively, in two thirds and one third of the bee venom-sensitive cases. Specific IgE antibody determinations by the Radioallergosorbent test play an essential role in the diagnostic work. After a reaction to hymenoptera stings both bee and wasp venom tests are necessary due to the high incidence of a false or incomplete identification of the stinging insect. Melittin, known for its potent pharmacological activity and possibly responsible for most of the side effects in bee venom immunotherapy, can probably not be excluded from therapeutic venom preparations since IgE antibodies to the melittin preparation were detected in one third of the cases.

Topics: Bee Venoms; Cross Reactions; Electrophoresis, Agar Gel; Humans; Hymenoptera; Immunoglobulin E; Insect Bites and Stings; Melitten; Phospholipases; Phospholipases A; Radioallergosorbent Test; Venoms; Wasps