Cross-reactivity Of Venoms Research Articles

Commercial Antivenoms Exert Broad Paraspecific Immunological Binding and In Vitro Inhibition of Medically Important Bothrops Pit Viper Venoms.

Snakebite envenoming is a life threatening neglected tropical disease that represents a considerable public health concern in the tropics. Viperid snakes of the genus Bothrops are among those of greatest medical importance in Latin America, and they frequently cause severe systemic haemotoxicity and local tissue destructive effects in human victims. Although snakebite antivenoms can be effective therapeutics, their efficacy is undermined by venom toxin variation among snake species. In this study we investigated the extent of paraspecific venom cross-reactivity exhibited by three distinct anti-Bothrops antivenoms (Soro antibotrópico-crotálico, BothroFav and PoliVal-ICP) against seven different Bothrops pit viper venoms from across Latin America. We applied a range of in vitro assays to assess the immunological binding and recognition of venom toxins by the antivenoms and their inhibitory activities against specific venom functionalities. Our findings demonstrated that, despite some variations, the monovalent antivenom BothroFav and the polyvalent antivenoms Soro antibotrópico-crotálico and PoliVap-ICP exhibited extensive immunological recognition of the distinct toxins found in the different Bothrops venoms, with Soro antibotrópico-crotálico generally outperformed by the other two products. In vitro functional assays revealed outcomes largely consistent with the immunological binding data, with PoliVap-ICP and BothroFav exhibiting the greatest inhibitory potencies against procoagulant and fibrinogen-depleting venom activities, though Soro antibotrópico-crotálico exhibited potent inhibition of venom metalloproteinase activities. Overall, our findings demonstrate broad levels of antivenom paraspecificity, with in vitro immunological binding and functional inhibition often highly comparable between venoms used to manufacture the antivenoms and those from related species, even in the case of the monovalent antivenom BothroFav. Our findings suggest that the current clinical utility of these antivenoms could possibly be expanded to other parts of Latin America that currently suffer from a lack of specific snakebite therapies.

A Rapid and International Applicable Diagnostic Device for Cobra (Genus Naja) Snakebites.

Cobra snakes (genus Naja) are some of the most dangerous snake species in Asia and Africa, as their bites cause severe life-threatening respiratory failure and local tissue destruction, especially in the case of late diagnosis. The differential diagnosis of snakebite envenomation still mainly relies upon symptomatology, the patient’s description, and the experience of physicians. We have designed a rapid test, immunochromatographic test of cobra (ICT-Cobra), which obtained fair results in improving the diagnosis and treatment of Naja (N.) atra snakebites in Taiwan. In this study, we further investigated the feasibility of applying the kit for the detection of other cobra venoms based on the potential interspecies similarity. We firstly demonstrated the cross-reactivity between eight venoms of medically important cobra species and the rabbit anti-N. atra IgG that was used in ICT-Cobra by Western blotting and sandwich enzyme-linked immunosorbent assay. Then, ICT-Cobra was used to detect various concentrations of the eight venoms to elucidate its performance. Noticeable correlations between the cross-reactivity of venoms from genus Naja snakes and existing geographical characteristics were found. ICT-Cobra could detect venoms from other Asian cobras with variable detection limits comparable to those observed for N. atra, but the kit was less successful in the detection of venom from African cobras. The similar but slightly different venom components and the interaction between venom and rabbit anti-N. atra IgG led to variations in the detection limits. The transcontinental usage of ICT-Cobra might be possible due to the cross-reactivity of antibodies and similarities among the larger-sized proteins. This study showed that the close immunological relationships in the genus Naja could be used to develop a venom detection kit for the diagnosis of cobra envenomation in both Asian and African regions. Additional clinical studies and technical adjustments are still needed to improve the efficacy and broadening the application of ICT-Cobra in the future.

Genus Calliophis of Asiatic coral snakes: A deficiency of venom cross-reactivity and neutralization against seven regional elapid antivenoms

Venoms of Calliophis bivirgata and Calliophis intestinalis exhibited moderate binding activities toward Neuro Bivalent Antivenom (Taiwan) but not the other six elapid monovalent or bivalent antivenoms available in the region. All antivenoms failed to neutralize C. bivirgata venom lethality in mice. The findings indicate the need to validate antivenom cross-reactivity with in vivo cross-neutralization, and imply that distinct antigens of Calliophis venoms should be incorporated in the production of a pan-regional poly-specific antivenom.

Immunological cross-reactivity of venoms from some Egyptian snakes and scorpians

The immunological cross-reactivity among the Elapidae, Viperidae and Buthidae venoms was detected in dot blot and western blot assays and quantified in ELISA as percentage of absorbance of heterologous versus homologous reaction. Mutual cross-reactivity between the Elapidae and Viperidae venoms was recognized, reflecting high inter-family relationship whereas only weak crossreactions were observed between the scorpion and snake venoms. The anti-scorpion antivenom exceptionally binds a 30-35 kDa Cerastes cerastes venom component, revealing common antigenic components between scorpions and C. cerastes venom. Many of the moderate and weak cross reactivity in ELISA got unnoticed in the dot blottechnique. This work primarily describes the commonality among venoms from remote source animals which is potentially important in the designation of venom detection kits and also in exploring common allergens responsible for cross-sensitization of patients to heterologous bites or stings leading to immediate hypersensitivity reactions.Keywords : Snakes, Scorpions, Cross-reactivity, Dot blot, ELISA.

Structural biology of allergens from stinging and biting insects

Modern techniques in genomic and protein research are applied to the study of stinging and biting insect allergens. Three-dimensional structures of additional insect venom and salivary allergens have been determined. An approach to determining B-cell epitopes has been used for hyaluronidase. A number of new venom and salivary allergens have been characterized. The structures and significance of several insect allergens have been updated. Investigations continue into distinguishing venom crossreactivity from multiple sensitization. Further studies are clarifying the significance of carbohydrate epitopes. Genomic and proteomic techniques are being used in the investigation of proteins and peptides in insect venom and saliva. The nature of venom crossreactivity and the B-cell and T-cell epitope structures of insect venom and salivary allergens are beginning to be elucidated.

A new assay for the detection of Loxosceles species (brown recluse) spider venom

Study Objective: Dermal lesions from unrelated arthropod species and medical causes appear similar to Loxosceles species (brown recluse spider) bites. This may result in delayed diagnosis and treatment. We developed a sensitive Loxosceles species venom enzyme-linked immunosorbent assay (ELISA) and characterized the specificity of the assay by evaluating antigenic cross-reactivity from a variety of North American arthropod venoms. Methods: North American arthropod (14 spiders, 2 scorpions, and 1 bee) venoms were studied. Three venom amounts (diluted in 100 μL of ELISA buffer) were assayed: 16,000 ng, 2,000 ng, and 40 ng. The latter quantity was selected because this is the observed maximum amount of venom we detect when inoculating dermis with amounts likely to be deposited by a spider bite. The larger venom amounts are overwhelming quantities designed to test the limits of the assay for arthropod venom cross-reactivity. Similar amounts of Loxosceles species venom and bovine albumin served as positive and negative controls, respectively. Results: At the lowest amount of venom tested (40 ng), the ELISA detected only the Loxosceles species positive control. When 2,000 ng was assayed, only Scytodes fusca and Kukulcania hibernalis arachnid venoms (in addition to Loxosceles species) cross-reacted to the assay. Finally, at 16,000 ng, the ELISA assay modestly detected Diguetia canities, Heteropoda venatoria, Tegenaria agrestis, Plectreurys tristes, Dolomedes tenebrosus, and Hadrurus arizonensis arachnid venoms. Conclusion: Cross-reactivity was observed in 8 of 17 North American arthropod venoms when large venom amounts were assayed with a Loxosceles species ELISA. By using a relevant quantity of venom, 40 ng, the assay was specific for Loxosceles species venom. The venom specificity of the ELISA may allow clinical application in Loxosceles species endemic regions of North America. [Gomez HF, Krywko DM, Stoecker WV. A new assay for the detection of Loxosceles species (brown recluse) spider venom. Ann Emerg Med. May 2002;39:469-474.]

Antigenic cross-reactivity of venoms from medically important North American Loxosceles spider species

We characterized the antigenic cross-reactivity of two medically important North American Loxoxceles species: L. reclusa (native to southeastern US) and L. deserta (native to southwestern US). Dermonecrosis resulting from bites from these two North American spider species are indistinguishable clinically. Polyclonal IgG antivenins directed against L. reclusa and L. deserta were raised in rabbits and used to develop specific enzyme immunoassays (EIAs). Antigenic differences in the two venoms were evaluated as follows: (1) Comparison of the sensitivities and correlation coefficient ( R 2) of anti- L. reclusa (αLoxR) and anti- L. deserta antibodies (αLoxD) in the detection of varying concentrations of the two venoms; (2) separation and western blot comparison of venom components; (3) protein sequence analysis of L. desertavenom and comparison to the L. reclusa protein sequence analysis present in a US national database; and (4) in vivo evaluation of αLoxR and αLoxD antivenins in attenuating dermal lesions (rabbit model). Correlation coefficients for αLoxR ( R 2=0.99) and αLoxD ( R 2=0.99) polyclonal antibodies in the measurements of standard concentrations of venoms were virtually identical. Western blot analysis revealed multiple common bands between the two venoms. Amino acid data (amino acids 1–35, N-terminal) of the active venom components of the two venoms revealed only three non-identical amino acids. αLoxR and αLoxD antivenins were similarly effective in blocking the development of rabbit skin lesions (ANOVA p<0.05). In summary, L. reclusa and L deserta spider venoms possess several common protein bands as identified by western blot, greater than 90% amino acid sequence identity, and marked antigenic cross-reactivity.

Antigenic cross-reactivity of venoms from medically important Loxosceles (araneae) species in Brazil

Antigenic cross-reactivity between the components of venoms from three spiders of the genus Loxosceles, L. gaucho, L. laeta and L. intermedia, was studied. Species-specific antisera were prepared by immunization of rabbits with each venom. Anti- L. gaucho horse hyperimmune serum provided by the Butantan Institute for treatment of accidents with these spiders was also used. Separation by SDS-PAGE showed the existence of many common components in the three antigens. No individual antigen was observed. Analysis of the antisera by ELISA and Western blotting showed cross-reactivity as well as several common bands between the three venoms. The horse anti- L. gaucho venom serum recognized many common proteins when antigens of the other two species were used. Antigens in the range of 33,000–35,000 mol. wt showed most cross-reactivity. Both horse and rabbit anti-venom sera contained antibodies able to neutralize the lethal and dermonecrotic activities of the venom of the three species studied.

Venom immunotherapy: 10 years of experience with administration of single venoms and 50 μg maintenance doses

For the past 10 years, we have administered venom immunotherapy with single venoms, whenever it is possible, and maintenance doses of 50 μg. The choice of venoms was based on clinical history, skin test reactions, and a knowledge of venom cross-reactivity. There have been. 258 re-stings in 108 patients with only three systemic reactions (2.7% per patient; 1.2% per sting). Two of these re-stings reactions were very mild, hives and facial edema, in patients who had had initial severe anaphylaxis. Five other patients had transient ill-defined symptoms, not considered allergic after re-stings. The patients covered a wide age range. Twenty-seven patients, nine under age 16 years, had initial dermal reactions only, and 44 patients had severe anaphylaxis. Most patients had multiple positive skin tests. Seventy-five patients received single venoms (yellow.jacket, 58; honeybee, 15; hornet, 2), and 30 patients received two venoms. Re-stings occurred from 1 month to 8 years, (mean, 2 years) after starting treatment. Results indicate that this approach with 50 μg top doses and single venom immunotherapy may be sufficient in most patients with an associated decrease in the cost as well as possible increased morbidity associated with the use of multiple venom antigens.

INSECT STING ALLERGY

Venom immunotherapy is highly effective prophylaxis for individuals at risk for insect sting anaphylaxis. Venom immunotherapy should be administered to all patients who have had a serious anaphylactic sting reaction (e.g., throat edema, dyspnea, hypotension, shock, loss of consciousness) and have positive venom skin tests. Children who have dermal (hives, edema) reactions only have a very benign course and do not require venom immunotherapy. Adults with similar mild sting reactions also appear to have a benign course, but data are still insufficient to recommend withholding venom immunotherapy. A positive venom skin test has been defined as a reaction to venom concentrations of 1.0 μg/mL or less. Reactions to only this top concentration of 1.0 μg/mL should be interpreted with caution, as nonspecific reactions may occur with this dose. Venom immunotherapy is usually administered with all venoms to which there is a positive skin test. Immunologic studies suggest that these multiple positive reactions are due to venom cross-reactivity. If the culprit insect can be identified, single venom therapy is effective. Recommended maintenance doses are 100 μg of single venoms and 300 μg of mixed vespid venoms. A lower dose, 50 μg of single venoms, appears to be adequate. After 1 year of therapy, the 4-week interval between maintenance doses can be extended to 6 and even 8 weeks, without loss of effectiveness or increased reactions. Recommendations for duration of therapy are evolving. Venom immunotherapy can be stopped if skin test reactions become negative. For most patients, 3 years of therapy appears adequate, despite persistence of positive venom skin tests.

Antigenic cross-reactivity of venoms obtained from snakes of genus Bothrops

Antigenic cross-reactivity of venoms obtained from snakes of genus Bothrops. Toxicon 28, 181–188, 1990.—Antigenic cross-reactivity was studied among the components of venoms from nine species of the genus Bothrops using species-specific antivenoms. Sera titration by DOT-ELISA detected similar levels of antibody when either homologous or heterologous antigens were used. Transblotted antigens, after SDS-PAGE fractionation, were also revealed by homologous and heterologous antivenoms. Antigens with mol. wt greater than 30,000 seemed to be the most cross-reactive. Antigens of about 24,000 mol. wt were poorly immunogenic. Antigens between 14–18,000 mol. wt cross-reacted only with B. moojeni, B. jararacussu, B. neuwiedi and B. pradoi venoms. Neutralization of the lethality of B. jararaca venom was observed by homologous and heterologous antivenoms.

Polistes wasp hypersensitivity: Diagnosis by venom-induced release of histamine in vitro

Polistes wasps cause a majority of Hymenoptera-induced anaphylactic reactions in Texas. Using the in vitro release of histamine from basophils of patients allergic to Polistes stings, we have studied the cross-reactivity of venoms from three species of Polistes wasps as well as the cross-reactivity among Polistes, honeybee, and Vespula maculifrons (yellow jacket) venoms. Venom collected by an extrusion technique from Pollistes exclamans, Pollistes apachus, and Pollistes Carolina caused release of histamine in seven Polistes-sensitive individuals. The dose-response curves from all three Polistes species were quite similar, suggesting extensive cross-reactivity among these species. None of these patients showed significant release of histamine from leukocytes exposed to yellow jacket or honeybee venom. We conclude that a source of Polistes venom is available for further study and possibly for therapy. It appears that any of three local common species of Polistes wasps could be used. Our studies confirmed earlier reports that Hymenoptera sensitivity is often genus-specific.