Social Wasp Polybia Paulista Research Articles

SYNTHESIS AND BIOLOGICAL COMPARISON OF TWO ANTIMICROBIAL PEPTIDES ORIGINATED FROM THE VENOM OF VESPA CRABRO AND POLYBIA PAULISTA

Bee venom is abundant in potential antimicrobial peptides, AMPs, . Among them, Mastoparan C and Polybia-MP1 are two well-known AMPs that were found in the European Hornet Vespa crabro and social wasp Polybia paulista, respectively. Due to the potent antimicrobial activity toward a wide range of microbial pathogens, including Gram-negative, Gram-positive and fungal species, they are promising to tackle the global antimicrobial resistance crisis. Thus, understanding the structure and activity relationships (SARs) is essential for the development of potential therapeutic applications. In this study, we synthesized and compared the differences in biological properties of these two antimicrobial peptides. The preliminary data suggests various important information regarding the structural requirements for optimizing the pharmacological properties of antimicrobial peptides.

Cytotoxicity of polybia paulista wasp crude venom on the tumor cell lines HeLa, HRT-18, jurkat, and K562

The social wasp Polybia paulista is an animal that produces venom for defense, which has been the subject of numerous studies in recent decades. Despite the great scientific interest, its pharmacological and biotechnological potential is still unknown. In this study, the antitumor capacity of the crude venom of P. paulista was investigated through cytotoxicity assays on the cell lines HeLa, HRT-18, Jurkat, and K562. Based on the cytotoxicity assays performed after 24 hours of treatment, the results showed that the crude venom was able to significantly reduce cell proliferation and induce cell death in the K562, HRT-18, and Jurkat cell lines. Morphological and cell cycle analyses in the Jurkat cell line revealed evidence of chromatin fragmentation and the presence of early and late apoptotic cells. It can be concluded that the crude venom of the social wasp P. paulista is cytotoxic to the tumor cell lines K562 (chronic myeloid leukemia), Jurkat (acute lymphocytic leukemia), and HRT-18 (intestinal carcinoma). The obtained results report the cytotoxic effects of the venom of the wasp P. paulista and demonstrate the importance of prospecting this venom, which is still poorly investigated.

Effects of chronic exposure to sublethal doses of neonicotinoids in the social wasp Polybia paulista: Survival, mobility, and histopathology

Several studies have investigated the consequences of exposure to neonicotinoids in honeybees. Given the lack of studies concerning the consequences of exposure of social wasps to neonicotinoids, as well as the ecological importance of these insects, the objective of this study was to test the hypothesis that chronic exposure to sublethal concentrations of thiamethoxam decreases survival and mobility by causing damage to the brain and midgut of the social wasp Polybia paulista. The wasps were exposed to different concentrations of thiamethoxam, in order to obtain the mean lethal concentration (LC50), which was used as a reference for calculation of two sublethal concentrations (LC50/100 and LC50/10) employed in subsequent experiments. To calculate survival, groups of exposed (EW) and unexposed (UW) wasps were monitored until death, allowing calculation of the average lethal time. The EW and UW groups were evaluated after 12, 24, 48, and 72 h of exposure, considering their mobility and histopathological parameters of the midgut and brain. A lesion index based on semiquantitative analyses was used for comparison of histopathological damage. The results demonstrated that exposure to the LC50/10 led to a significantly shorter survival time of the P. paulista workers, compared to unexposed wasps. In addition, both sublethal concentrations decreased mobility and caused damage to the intestine (loss of brush border, presence of spherocrystals, loss of cytoplasmic material, and pyknosis) and the brain (loss of cell contact and pyknosis), regardless of the exposure time. The findings showed that, like bees, social wasps are nontarget insects susceptible to the detrimental consequences of neonicotinoid use, with exposure leading to impaired survival, locomotion, and physiology.

Is the social wasp Polybia paulista a silent victim of neonicotinoid contamination?

Neonicotinoids are among the chemicals most widely used against insects considered agricultural pests, although they may also affect nontarget species, as has been reported for social bees. Social wasps are recognized as efficient predators of larvae of other insects, including pest species, so they may have contact with insecticides, at least indirectly. However, to date, there have been no studies investigating the consequences for social wasps of the use of neonicotinoids. Therefore, the aim of this study was to obtain the topical and oral acute lethal mean doses of the neonicotinoid thiamethoxam, as well as to evaluate the effects of sublethal contamination, for the social wasp Polybia paulista. Foraging wasps were exposed orally and topically, with monitoring of their mortality after 24 and 48 h, in order to calculate the LD50 values. Other wasps were then exposed with sublethal doses, in order to evaluate the effects of contamination on their mobility. The results indicated that P. paulista workers are as sensitive to thiamethoxam contamination as several nontarget species studied previously. Exposure to a sublethal dose (LD10) negatively affected their mobility. It could be concluded that social wasp colonies may be as much affected by exposure to neonicotinoids as social bees. Further toxicological studies should be conducted, evaluating other parameters and different species, in order to understand the extent of the problems faced by these insects due to the use of neonicotinoids.

Biological and Inflammatory Effects of Antigen 5 from Polybia paulista (Hymenoptera, Vespidae) Venom in Mouse Intraperitoneal Macrophages.

The social wasp Polybia paulista (Hymenoptera, Vespidae) is highly aggressive, being responsible for many medical occurrences. One of the most allergenic components of this venom is Antigen 5 (Poly p 5). The possible modulation of the in vitro immune response induced by antigen 5 from P. paulista venom, expressed recombinantly (rPoly p 5), on BALB/c mice peritoneal macrophages, activated or not with LPS, was assessed. Here, we analyzed cell viability changes, expression of the phosphorylated form of p65 NF-κB subunit, nitric oxide (NO), proinflammatory cytokines production, and co-stimulatory molecules (CD80, CD86). The results suggest that rPoly p 5 does not affect NO production nor the expression of co-stimulatory molecules in mouse peritoneal macrophages. On the other hand, rPoly p 5 induced an increase in IL-1β production in non-activated macrophages and a reduction in the production of TNF-α and MCP-1 cytokines in activated macrophages. rPoly p 5 decreased the in vitro production of the phosphorylated p65 NF-κB subunit in non-activated macrophages. These findings suggest an essential role of this allergen in the polarization of functional M2 macrophage phenotypes, when analyzed in previously activated macrophages. Further investigations, mainly in in vivo studies, should be conducted to elucidate Polybia paulista Ag5 biological role in the macrophage functional profile modulation.

The effects of incorporation of the counterparts and mimics of L-lysine on the antimicrobial activity, hemolytic activity, cytotoxicity and tryptic stability of antimicrobial peptide polybia-MPII.

Due to the limited effects of conventional antibiotics on the increasing emergence of drug-resistant bacteria and fungi, novel antimicrobial agents were urgently needed to alleviate this phenomenon. Nowadays, antimicrobial peptides are believed to be a promising candidate for a new generation of antimicrobial drugs. Antimicrobial peptide polybia-MPII (MPII) was first isolated from the venom of the social wasp Polybia paulista with a broad spectrum of antimicrobial activity. In the present study, the counterparts and mimics of cationic amino acids of Lys, such as Arg, His, Orn, Dab and Dap were employed to substitute Lys in the sequence of MPII. The effects of the incorporation of these amino acids on its antimicrobial activity, hemolytic activity, cytotoxicity, enzyme stability and therapeutic potential were explored. Our results showed that although the incorporation of Arg could improve its antimicrobial activity, there is no improvement in enzyme stability. The incorporation of His makes MPII exert its antimicrobial activity in a pH-dependent manner. Notably, incorporating Dap could effectively decrease its hemolytic activity and cytotoxicity and enhance its enzyme stability against trypsin. In conclusion, this study would provide an effective strategy to improve the bioavailability and metabolic stability of AMPs while decrease their hemolytic activity and cytotoxicity.

Is It Possible to Obtain the Chemical Profile From Ethanol-Preserved Specimens? The Hydrocarbon and Fatty Acid Composition of the Social Wasp Polybia paulista (Hymenoptera: Vespidae: Epiponini).

Cuticular hydrocarbons perform multiple functions in insects such as protecting against desiccation and pathogenic infection, and signaling interactions. Evaluation of cuticular hydrocarbon (CHC) profiles of insects is commonly performed by extraction using a nonpolar solvent such as hexane. Specimens intended for CHC analysis are ideally handled by avoiding contact with solvents such as ethanol. However, insects are frequently stored in ethanol after collection, especially if intended for molecular analysis. To determine if it is possible that chemical compounds in the cuticles of specimens can withstand previous exposure to ethanol, we evaluated the efficiency of CHC extraction from specimens preserved in 95% ethanol. We extracted cuticular compounds from specimens of the social wasp Polybia paulista (Ihering) with no contact with ethanol solvents and compared them with those from specimens stored in 95% ethanol. We analyzed chemical composition from wasps and the 95% ethanol in which they had been stored by a gas chromatograph coupled to a mass spectrometer. In total, 56 compounds were detected: 50 that were classified as hydrocarbons which were mostly branched alkanes, followed by linear alkanes and alkenes. Three compounds were identified as fatty acids, and three compounds were unidentifiable. The ethanol-preserved specimens showed similar chemical profiles to those of specimens that had no contact with ethanol. Thus, we suggest that it is possible to study the chemical profiles of ethanol-preserved specimens.

Neuropolybin: A new antiseizure peptide obtained from wasp venom

Epilepsy accounts for one of the most serious neurological disorders, and its treatment remains a challenge, due to high cost and harmful side effects. Bioactive molecules extracted from arthropod venoms are considered a promising therapy since these compounds are known for their highly selective and potent profiles. The purpose of this study was to identify and characterize the potential antiseizure effect of the peptide Ppnp7, extracted from the venom of the social wasp Polybia paulista, and also the effect of the bioinspired peptide, named Neuropolybin, in the same parameters. Additionally, we also evaluated the electroencephalographic (EEG) profile in the PTZ-induced acute seizures in animals treated with Neuropolybin, and potential adverse effects of both peptides in general spontaneous activity (Open Field analysis). Interestingly, Ppnp7 and Neuropolybin showed a noteworthy antiseizure effect in rats and mice, respectively. Curves of protection against the maximum seizure were obtained for both peptides, and EEG records demonstrated that Neuropolybin protected 80% of animals from tonic-clonic seizures when applied with a dose of 3 nmol (an approximate Ppnp7 ED50 found in rats). Neuropolybin and Ppnp7 did not cause changes in the general spontaneous activity of the animals in any of the doses evaluated. Therefore, this study demonstrated how compounds isolated from wasps’ venom may be essential resources in the search for new drugs, and can also be considered valuable therapeutic and biotechnological tools for the study and future treatment of epileptic disorders.

Evaluation of Inter and Intraspecific Differences in the Venom Chemical Compositions of Polybia paulista Wasps and Ectatomma brunneum Ants Using FTIR-PAS

Wasps can synthesize chemical compounds called venom whose function is to overcome prey and assist in defense of the colonies. Geographic Parameters such as sex, age, the season of the year, and diet determined the composition of the venom location, genetics, environment. However, studies on the compositional variability of venom are still limited due to the difficulty in obtaining samples and the complexity of these substances. This work describes the use of the Fourier Transform Infrared Photoacoustic Spectroscopy (FTIR-PAS) to investigate inter- and intraspecific variability in the venom chemical composition (VCC) of the social wasp Polybia paulista (Von Ihering 1896) and the ant Ectatomma brunneum (Smith 1858). The results reveal significant differences in VCC among the ant and wasp, even for samples obtained from the same environment. The genetic component, therefore, seemed to be the predominant factor determining the compounds present. The findings also showed that exogenous factors, such as diet, could also be responsible for intraspecific differences, especially in wasps. The FTIR-PAS technique proved to be a reliable way of assessing intra- and interspecific differences in social Hymenoptera VCC.

Effect of Seasonality on Rates of Gregarine Infection in Workers of a Social Wasp Polybia Paulista (Hymenoptera, Vespidae).

Social insects face strong selection from parasites because the conditions of group living often favor the transmission of infection among nestmates. However, there is little detailed information on the effects of parasite infection in the host species. Workers of Polybia species, neotropical swarm-founding wasps, are commonly infected by gregarines, protozoans that are exclusively parasitic on invertebrates. Previous studies showed that high rates of gregarine infection in workers of Polybia occidentalis (Olivier) have negative effects on their colony performance. However, the effect of seasonality on infection rates throughout the year or between wet and dry seasons has not been examined. Host-parasite interactions cannot be understood without consideration of the overall population dynamic. We compared rates of gregarine infection in workers of Polybia paulista (Ihering) between wet and dry seasons and among months. The 35% rate was by far the highest of the four wet seasons sampled, but the rates declined in the mid-wet season and were very low during the dry season. Strong seasonal differences in infection rates were also observed between the dry and wet seasons. Several potential factors affecting the seasonal differences are discussed.

Antimicrobial activity and stability of stapled helices of polybia-MP1.

Polybia-MP1 is a well-known natural antimicrobial peptide isolated from the venom of the social wasp Polybia paulista. A recent study showed that this peptide displays a broad antibacterial spectrum as well as low toxicity to human red blood cells and normal fibroblasts. However, its moderate antimicrobial activity and high susceptibility to protease have been a major hurdle for clinical use. This study examined the possibility of developing biologically more potent, yet metabolically more stable, analogues of MP1 using an emerging technology termed "all-hydrocarbon stapling." The stapled analogues of MP1 showed more than a threefold increase in helicity as well as an approximately 70-fold enhancement in proteolytic stability. These stapled analogues also exhibited a significant increase in inhibition against some Gram-positive bacteria while displaying a modest enhancement in hemolytic activity. Overall, the current study demonstrated that the all-hydrocarbon stapling system is a highly useful tool for the development of biologically more potent and metabolically more stable analogues of natural antimicrobial peptides.

Anxiolytic activity and evaluation of potentially adverse effects of a bradykinin-related peptide isolated from a social wasp venom

Anxiety disorders are major health problems in terms of costs stemming from sick leave, disabilities, healthcare and premature mortality. Despite the availability of classic anxiolytics, some anxiety disorders are still resistant to treatment, with higher rates of adverse effects. In this respect, several toxins isolated from arthropod venoms are useful in identifying new compounds to treat neurological disorders, particularly pathological anxiety. Thus, the aims of this study were to identify and characterize an anxiolytic peptide isolated from the venom of the social wasp Polybia paulista. The peptide was identified as Polisteskinin R, with nominal molecular mass [M+H]+=1301Da and primary structure consisting of Ala-Arg-Arg-Pro-Pro-Gly-Phe-Thr-Pro-Phe-Arg-OH. The anxiolytic effect was tested using the elevated plus maze test. Moreover, adverse effects on the spontaneous behavior and motor coordination of animals were assessed using the open field and rotarod tests. Polisteskinin R induced a dose-dependent anxiolytic effect. Animals treated with the peptide and diazepam spent significantly more time into the open arms when compared to the groups treated with the vehicle and pentylenetetrazole. No significant differences in spontaneous behavior or motor coordination were observed between the groups, showing that the peptide was well tolerated. The interaction by agonists in both known BK receptors induces a variability of physiological effects; Polisteskinin R can act on these receptors, inducing modulatory activity and thus, attenuating anxiety behaviors. The results of this study demonstrated that the compound Polisteskinin R exerted potent anxiolytic effects and its analogues are promising candidates for experimental pharmacology.

Using a proteometabolomic approach to investigate the role of Dufour's gland in pheromone biosynthesis in the social wasp Polybia paulista

Dufour's gland is associated with the venom apparatuses of social wasps and bees. This location and its evolutionary adaptations indicate that it could be involved in the production of alarm pheromones in the social wasp Polybia paulista. To investigate this hypothesis, the volatile composition of this gland was analyzed and compared to that in the venom. Eighteen compounds were identified as secreted by Dufour's gland, and 16 of these compounds were also identified in the venom, suggesting that the compounds produced by the gland are secreted and mixed with venom in the venom reservoir of this wasp. These compounds were subjected to a field bioassay to investigate their potential action as alarm pheromones. Alcohols and aldehydes elicited the alert behavior in workers, luring them outside the nest, whereas acids attracted the workers in the direction of the source; fatty acid methyl esters elicited aggression. These results suggest that Dufour's gland produces alarm pheromones. To corroborate this hypothesis the proteomic complement of this gland was assigned using a shot-gun strategy; 59 proteins were identified, and the results indicate specialization of Dufour's gland for the metabolism of fatty acids (elongation, esterification unsaturation, reduction, and decarboxylation) in the biosynthesis of alarm pheromones. Biological significanceThe present knowledge about the role of Dufour's gland among aculeate Hymenoptera insects suggests that it may have many different roles related to the biosynthesis and secretion of chemical markers for different biological functions, though none are related to the elicitation of alarm behaviors for coordinating a mass attack of the colony against intruders. The present study combined the analysis of secreted volatile compounds (as metabolites) with proteome assignments and a field bioassay with synthetic compounds to clearly demonstrate that Dufour's gland does in fact biosynthesize alarm pheromones in social wasps. This strategy may be reproduced in other investigations related to pheromone production in other insects.

Antifungal effect and action mechanism of antimicrobial peptide polybia‐CP

The incidence of life-threatening invasive fungal infections increased significantly in recent years. However, the antifungal therapeutic options are very limited. Antimicrobial peptides are a class of potential lead chemical for the development of novel antifungal agents. Antimicrobial peptide polybia-CP was purified from the venom of the social wasp Polybia paulista. In this study, we synthesized polybia-CP and determined its antifungal effects against a series of Candidian species. Our results showed that polybia-CP has potent antifungal activity and fungicidal activity against the tested fungal cells with a proposed membrane-active action mode. In addition, polybia-CP could induce the increase of cellular reactive oxygen species production, which would attribute to its antifungal activity. In conclusion, the present study suggests that polybia-CP has potential as an antifungal agent or may offer a new strategy for antifungal therapeutic option.

Dual antifungal properties of cationic antimicrobial peptides polybia-MPI: Membrane integrity disruption and inhibition of biofilm formation

With the increasing emergence of resistant fungi, the discovery and development of novel antifungal therapeutics were urgently needed. Compared with conventional antibiotics, the limited propensity of AMPs to induce resistance in pathogens has attracted great interest. In the present study, the antifungal activity and its mechanism-of-action of polybia-MPI, a cationic peptide from the venom of Social wasp Polybia Paulista was investigated. We demonstrated that polybia-MPI could potently inhibit the growth of Candida albicans (C. albicans) and Candida glabrata (C. glabrata). The 50% inhibitory concentrations (IC50) of Polybia-MPI against cancer cells were much higher than the MICs against the tested C. albicans and C. glabrata cells, indicating that polybia-MPI had high selectivity between the fungal and mammalian cells. Our results also indicated that membrane disturbance mechanism was involved in the antifungal activity. Furthermore, polybia-MPI could inhibit the bio film forming of C. glabrata, which was frequently associated with clinically significant biofilm. These results suggest that polybia-MPI has great advantages in the development of antifungal agents.

Reactivity of IgE to the allergen hyaluronidase from Polybia paulista (Hymenoptera, Vespidae) venom

To date, there are no allergenic extracts or components available in Brazil to diagnosis and treatment of patients with venom allergy from social wasp (Vespidae Family; Polistinae Subfamily) despite of the great number of existing species. We evaluated the immunogenic potential of the Hyal recombinant protein (Pp-Hyal-rec) which was expressed in an insoluble form in comparison with the allergenic native protein (Pp-Hyal-nat) for recognition of immunoglobulin E (IgE) in the serum of allergic patients to venom of the endemic social wasp Polybia paulista from São Paulo State, Brazil. Hyal cDNA from the venom of the social wasp P. paulista (Pp-Hyal) (GI: 302201582) was cloned into the expression vector pET-28a in Escherichia coli DE3 (BL21) cells. Solubilization and purification of Pp-Hyal-rec from inclusion bodies were performed using Ni2+ affinity chromatography (Ni-NTA-Agarose) under denaturing conditions. Both the native (Pp-Hyal-nat) and the recombinant (Pp-Hyal-rec) purified allergens were used for Western blotting to assess the levels of Pp-Hyal-IgE specific in the serum of 10 patients exclusively reactive to the venom of the social wasp P. paulista. The immune sera specifically recognized the band corresponding to the Pp-Hyal-rec protein (40 kDa) at a higher intensity than the native allergen (39 kDa). The sera recognized other proteins in P. paulista crude venom extract to a lesser extent, likely corresponding to other venom allergens such as phospholipase (34 kDa), Antigen 5 (25 kDa), and proteases. The recognition pattern of the immune sera to the Pp-Hyal-rec allergen strongly suggests that this recombinant antigen could be used for developing a diagnostic allergy test as well as for specific immunotherapy (IT).

Using Proteomic Strategies for Sequencing and Post-Translational Modifications Assignment of Antigen-5, a Major Allergen from the Venom of the Social Wasp Polybia paulista

Antigen-5 is one of the major allergens identified in wasp venoms, and despite the fact that its biological function is still unknown, many studies have demonstrated its allergenicity. In this study, the biochemical and structural characterization of antigen-5 from the venom of the social wasp Polybia paulista are reported. A gel-based mass spectrometry strategy with CID fragmentation methods and classical protocols of protein chemistry, which included N- and C-terminal sequencing, were used to assign the complete sequence and determine the presence/location of the post-translational modifications (PTMs) of this protein. Six different isoforms of antigen-5 were identified in the crude venom of P. paulista ; the most abundant, which corresponds to the intact form of this protein, was recognized by the pool of human specific-IgE. This protein was extensively sequenced through CID mass spectrometry, and a series of PTMs were observed such as hydroxylation, phosphorylation, and glycosylation. Sequence data revealed that this protein has 59.3-93.7% identity with antigen-5 proteins from other known vespid venoms. The molecular model of P. paulista antigen-5 shows that this protein has three α-helices, one 310 helix, and four β-sheets covering 28 and 17.9% of the sequence, respectively. The identification and characterization of allergenic compounds is essential for the development of advanced component-resolved allergy diagnostics and treatment.

Peptide diversity in the venom of the social wasp Polybia paulista (Hymenoptera): A comparison of the intra- and inter-colony compositions

The venoms of the social wasps evolved to be used as defensive tools to protect the colonies of these insects against the attacks of predators. Previous studies estimated the presence of a dozen peptide components in the venoms of each species of these insects, which altogether comprise up to 70% of the weight of freeze-dried venoms. In the present study, an optimized experimental protocol is reported that utilizes liquid chromatography coupled to electrospray ionization mass spectrometry for the detection of peptides in the venom of the social wasp Polybia paulista; peptide profiles for both intra- and inter-colonial comparisons were obtained using this protocol. The results of our study revealed a surprisingly high level of intra- and inter-colonial variability for the same wasp species. We detected 78–108 different peptides in the venom of different colonies of P. paulista in the molar mass range from 400 to 3000Da; among those, only 36 and 44 common peptides were observed in the inter- and intra-colony comparisons, respectively.

Structure–function relationships of the peptide Paulistine: A novel toxin from the venom of the social wasp Polybia paulista

BackgroundThe peptide Paulistine was isolated from the venom of wasp Polybia paulista. This peptide exists under a natural equilibrium between the forms: oxidised — with an intra-molecular disulphide bridge; and reduced — in which the thiol groups of the cysteine residues do not form the disulphide bridge. The biological activities of both forms of the peptide are unknown up to now. MethodsBoth forms of Paulistine were synthesised and the thiol groups of the reduced form were protected with the acetamidemethyl group [Acm-Paulistine] to prevent re-oxidation. The structure/activity relationships of the two forms were investigated, taking into account the importance of the disulphide bridge. ResultsPaulistine has a more compact structure, while Acm-Paulistine has a more expanded conformation. Bioassays reported that Paulistine caused hyperalgesia by interacting with the receptors of lipid mediators involved in the cyclooxygenase type II pathway, while Acm-Paullistine also caused hyperalgesia, but mediated by receptors involved in the participation of prostanoids in the cyclooxygenase type II pathway. ConclusionThe acetamidemethylation of the thiol groups of cysteine residues caused small structural changes, which in turn may have affected some physicochemical properties of the Paulistine. Thus, the dissociation of the hyperalgesy from the edematogenic effect when the actions of Paulistine and Acm-Paulistine are compared to each other may be resulting from the influence of the introduction of Acm-group in the structure of Paulistine. General significanceThe peptides Paulistine and Acm-Paulistine may be used as interesting tools to investigate the mechanisms of pain and inflammation in future studies.

Hyaluronidase from the venom of the social wasp Polybia paulista (Hymenoptera, Vespidae): Cloning, structural modeling, purification, and immunological analysis

In this study, we describe the cDNA cloning, sequencing, and 3-D structure of the allergen hyaluronidase from Polybia paulista venom (Pp-Hyal). Using a proteomic approach, the native form of Pp-Hyal was purified to homogeneity and used to produce a Pp-specific polyclonal antibody. The results revealed that Pp-Hyal can be classified as a glycosyl hydrolase and that the full-length Pp-Hyal cDNA (1315 bp; GI: 302201582) is similar (80–90%) to hyaluronidase from the venoms of endemic Northern wasp species. The isolated mature protein is comprised of 338 amino acids, with a theoretical pI of 8.77 and a molecular mass of 39,648.8 Da versus a pI of 8.13 and 43,277.0 Da indicated by MS. The Pp-Hyal 3D-structural model revealed a central core (α/β)7 barrel, two sulfide bonds (Cys 19–308 and Cys 185–197), and three putative glycosylation sites (Asn79, Asn187, and Asn325), two of which are also found in the rVes v 2 protein. Based on the model, residues Ser299, Asp107, and Glu109 interact with the substrate and potential epitopes (five conformational and seven linear) located at surface-exposed regions of the structure. Purified native Pp-Hyal showed high similarity (97%) with hyaluronidase from Polistes annularis venom (Q9U6V9). Immunoblotting analysis confirmed the specificity of the Pp-Hyal-specific antibody as it recognized the Pp-Hyal protein in both the purified fraction and P. paulista crude venom. No reaction was observed with the venoms of Apis mellifera, Solenopsis invicta, Agelaia pallipes pallipes, and Polistes lanio lanio, with the exception of immune cross-reactivity with venoms of the genus Polybia (sericea and ignobilis). Our results demonstrate cross-reactivity only between wasp venoms from the genus Polybia. The absence of cross-reactivity between the venoms of wasps and bees observed here is important because it allows identification of the insect responsible for sensitization, or at least of the phylogenetically closest insect, in order to facilitate effective immunotherapy in allergic patients.