Antiradiation Antitoxin IgG : Immunological neutralization of Radiation Toxins at Acute Radiation Syndromes.

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Popov, Dmitri; Maliev, Slava

Introduction: High doses of radiation induce apoptotic necrosis of radio-sensitive cells. Mild doses of radiation induce apoptosis or controlled programmed death of radio-sensitive cells with-out development of inflammation and formation of Radiation Toxins. Cell apoptotic necrosis initiates Radiation Toxins (RT)formation. Radiation Toxins play an important role as a trig-ger mechanism for inflammation development and cell lysis. If an immunotherapy approach to treatment of the acute radiation syndromes (ARS) were to be developed, a consideration could be given to neutralization of radiation toxins (Specific Radiation Determinants-SRD) by specific antiradiation antibodies. Therapeutic neutralization effects of the blocking anti-radiation antibodies on the circulated RT had been studied. Radiation Toxins were isolated from the central lymph of irradiated animals with Cerebrovascular(Cv ARS),Cardiovascular (Cr ARS),Gastrointestinal(Gi ARS) and Haemopoietic (Hp ARS) forms of ARS. To accomplish this objective, irradiated animals were injected with a preparation of anti-radiation immunoglobulin G (IgG) obtained from hyperimmune donors. Radiation-induced toxins that we call Specific Radiation Determinants (SRD) possess toxic (neurotoxic, haemotoxic) characteristics as well as specific antigenic properties. Depending on direct physiochemical radiation damage, they can induce development of many of the pathological processes associated with ARS. We have tested several specific hyperimmune IgG preparations against these radiation toxins and ob-served that their toxic properties were neutralized by the specific antiradiation IgGs. Material and Methods: A scheme of experiments was following: 1.Isolation of radiation toxins (RT) from the central lymph of irradiated animals with different form of ARS. 2.Transformation of a toxic form of the RT to a toxoid form of the RT. 3.Immunization of radiation naive animals. Four groups of rabbits were inoculated with a toxoid form of SRD

Mapping the epitopes of a neutralizing antibody fragment directed against the lethal factor of Bacillus anthracis and cross-reacting with the homologous edema factor.

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Philippe Thullier

Full Text Available The lethal toxin (LT of Bacillus anthracis, composed of the protective antigen (PA and the lethal factor (LF, plays an essential role in anthrax pathogenesis. PA also interacts with the edema factor (EF, 20% identity with LF to form the edema toxin (ET, which has a lesser role in anthrax pathogenesis. The first recombinant antibody fragment directed against LF was scFv 2LF; it neutralizes LT by blocking the interaction between PA and LF. Here, we report that scFv 2LF cross-reacts with EF and cross-neutralizes ET, and we present an in silico method taking advantage of this cross-reactivity to map the epitope of scFv 2LF on both LF and EF. This method identified five epitope candidates on LF, constituted of a total of 32 residues, which were tested experimentally by mutating the residues to alanine. This combined approach precisely identified the epitope of scFv 2LF on LF as five residues (H229, R230, Q234, L235 and Y236, of which three were missed by the consensus epitope candidate identified by pre-existing in silico methods. The homolog of this epitope on EF (H253, R254, E258, L259 and Y260 was experimentally confirmed to constitute the epitope of scFv 2LF on EF. Other inhibitors, including synthetic molecules, could be used to target these epitopes for therapeutic purposes. The in silico method presented here may be of more general interest.

Determination of low tetanus or diphtheria antitoxin titers in sera by a toxin neutralization assay and a modified toxin-binding inhibition test

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M.H. Sonobe

2007-01-01

Full Text Available A method for the screening of tetanus and diphtheria antibodies in serum using anatoxin (inactivated toxin instead of toxin was developed as an alternative to the in vivo toxin neutralization assay based on the toxin-binding inhibition test (TOBI test. In this study, the serum titers (values between 1.0 and 19.5 IU measured by a modified TOBI test (Modi-TOBI test and toxin neutralization assays were correlated (P < 0.0001. Titers of tetanus or diphtheria antibodies were evaluated in serum samples from guinea pigs immunized with tetanus toxoid, diphtheria-tetanus or triple vaccine. For the Modi-TOBI test, after blocking the microtiter plates, standard tetanus or diphtheria antitoxin and different concentrations of guinea pig sera were incubated with the respective anatoxin. Twelve hours later, these samples were transferred to a plate previously coated with tetanus or diphtheria antitoxin to bind the remaining anatoxin. The anatoxin was then detected using a peroxidase-labeled tetanus or diphtheria antitoxin. Serum titers were calculated using a linear regression plot of the results for the corresponding standard antitoxin. For the toxin neutralization assay, L+/10/50 doses of either toxin combined with different concentrations of serum samples were inoculated into mice for anti-tetanus detection, or in guinea pigs for anti-diphtheria detection. Both assays were suitable for determining wide ranges of antitoxin levels. The linear regression plots showed high correlation coefficients for tetanus (r² = 0.95, P < 0.0001 and for diphtheria (r² = 0.93, P < 0.0001 between the in vitro and the in vivo assays. The standardized method is appropriate for evaluating titers of neutralizing antibodies, thus permitting the in vitro control of serum antitoxin levels.

Bacillus anthracis lethal toxin disrupts TCR signaling in CD1d-restricted NKT cells leading to functional anergy.

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Sunil K Joshi

2009-09-01

Full Text Available Exogenous CD1d-binding glycolipid (alpha-Galactosylceramide, alpha-GC stimulates TCR signaling and activation of type-1 natural killer-like T (NKT cells. Activated NKT cells play a central role in the regulation of adaptive and protective immune responses against pathogens and tumors. In the present study, we tested the effect of Bacillus anthracis lethal toxin (LT on NKT cells both in vivo and in vitro. LT is a binary toxin known to suppress host immune responses during anthrax disease and intoxicates cells by protective antigen (PA-mediated intracellular delivery of lethal factor (LF, a potent metalloprotease. We observed that NKT cells expressed anthrax toxin receptors (CMG-2 and TEM-8 and bound more PA than other immune cell types. A sub-lethal dose of LT administered in vivo in C57BL/6 mice decreased expression of the activation receptor NKG2D by NKT cells but not by NK cells. The in vivo administration of LT led to decreased TCR-induced cytokine secretion but did not affect TCR expression. Further analysis revealed LT-dependent inhibition of TCR-stimulated MAP kinase signaling in NKT cells attributable to LT cleavage of the MAP kinase kinase MEK-2. We propose that Bacillus anthracis-derived LT causes a novel form of functional anergy in NKT cells and therefore has potential for contributing to immune evasion by the pathogen.

EL4 cell-based colorimetric toxin neutralization activity assays for determination of neutralizing anti-ricin antibodies.

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Lindsey, Changhong Y; Brown, J Edward; Torabazar, Nahid R; Smith, Leonard A

2013-01-01

A recombinant ricin toxin A-chain 1-33/44-198 vaccine (RVEc), developed at the United States Army Medical Research Institute of Infectious Diseases as a vaccine candidate, is under investigation in a phase 1 clinical study. To effectively evaluate the immunogenicity of this ricin vaccine and to eliminate the use of radioactive material, an EL4 cell-based colorimetric toxin neutralization activity (TNA) assay using a CellTiter 96 AQueous One Solution Cell Proliferation Assay Reagent has been developed, optimized, and applied in the vaccine efficacy studies. The TNA assay measures the protective neutralizing anti-ricin antibodies in animal sera by determining the cell viability after ricin exposure in the assay system and comparing it to a purified mouse polyclonal antiricin IgG standard curve. The standard curve of the anti-ricin TNA assay closely fits a four-parameter logistic regression model. The unknown test sample concentration was expressed as microg/mL, but not the 50% effective concentration (EC50), which was determined by most TNA assays. The neutralizing endpoint titers, not the 50% effective dilution (ED50), of human specimens were measured with the TNA assay in support of the clinical study of the RVEc vaccine. The optimal amount of ricin toxin, EL4 cells, and concentration of standards used in the assay system was established to minimize false-negative and false-positive results of serum specimens from the nonclinical and clinical studies of RVEc. The testing conditions were adjusted to optimize assay performance. The colorimetric TNA assay replaced a radioactive TNA assay previously used in the ricin vaccine studies.

Revisiting Notechis scutatus venom: on shotgun proteomics and neutralization by the "bivalent" Sea Snake Antivenom.

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Tan, Choo Hock; Tan, Kae Yi; Tan, Nget Hong

2016-07-20

Recent advances in proteomics enable deep profiling of the compositional details of snake venoms for improved understanding on envenomation pathophysiology and immunological neutralization. In this study, the venom of Australian tiger snake (Notechis scutatus) was trypsin-digested in solution and subjected to nano-ESI-LCMS/MS. Applying a relative quantitative proteomic approach, the findings revealed a proteome comprising 42 toxin subtypes clustered into 12 protein families. Phospholipases A2 constitute the most abundant toxins (74.5% of total venom proteins) followed by Kunitz serine protease inhibitors (6.9%), snake venom serine proteases (5.9%), alpha-neurotoxins (5.6%) and several toxins of lower abundance. The proteome correlates with N. scutatus envenoming effects including pre-synaptic and post-synaptic neurotoxicity and consumptive coagulopathy. The venom is highly lethal in mice (intravenous median lethal dose=0.09μg/g). BioCSL Sea Snake Antivenom, raised against the venoms of beaked sea snake (Hydrophis schistosus) and N. scutatus (added for enhanced immunogenicity), neutralized the lethal effect of N. scutatus venom (potency=2.95mg/ml) much more effectively than the targeted H.schistosus venom (potency=0.48mg/ml). The combined venom immunogen may have improved the neutralization against phospholipases A2 which are abundant in both venoms, but not short-neurotoxins which are predominant only in H. schistosus venom. A shotgun proteomic approach adopted in this study revealed the compositional details of the venom of common tiger snake from Australia, Notechis scutatus. The proteomic findings provided additional information on the relative abundances of toxins and the detection of proteins of minor expression unreported previously. The potent lethal effect of the venom was neutralized by bioCSL Sea Snake Antivenom, an anticipated finding due to the fact that the Sea Snake Antivenom is actually bivalent in nature, being raised against a mix of venoms of the

Neutralization of Clostridium difficile Toxin B Mediated by Engineered Lactobacilli That Produce Single-Domain Antibodies

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Andersen, Kasper Krogh; Strokappe, Nika M.; Hultberg, Anna; Truusalu, Kai; Smidt, Imbi; Mikelsaar, Raik-Hiio; Mikelsaar, Marika; Verrips, Theo; Hammarström, Lennart

2015-01-01

Clostridium difficile is the primary cause of nosocomial antibiotic-associated diarrhea in the Western world. The major virulence factors of C. difficile are two exotoxins, toxin A (TcdA) and toxin B (TcdB), which cause extensive colonic inflammation and epithelial damage manifested by episodes of diarrhea. In this study, we explored the basis for an oral antitoxin strategy based on engineered Lactobacillus strains expressing TcdB-neutralizing antibody fragments in the gastrointestinal tract. Variable domain of heavy chain-only (VHH) antibodies were raised in llamas by immunization with the complete TcdB toxin. Four unique VHH fragments neutralizing TcdB in vitro were isolated. When these VHH fragments were expressed in either secreted or cell wall-anchored form in Lactobacillus paracasei BL23, they were able to neutralize the cytotoxic effect of the toxin in an in vitro cell-based assay. Prophylactic treatment with a combination of two strains of engineered L. paracasei BL23 expressing two neutralizing anti-TcdB VHH fragments (VHH-B2 and VHH-G3) delayed killing in a hamster protection model where the animals were challenged with spores of a TcdA− TcdB+ strain of C. difficile (P survived until the termination of the experiment at day 5 and showed either no damage or limited inflammation of the colonic mucosa despite having been colonized with C. difficile for up to 4 days. The protective effect in the hamster model suggests that the strategy could be explored as a supplement to existing therapies for patients. PMID:26573738

SiMa Cells for a Serotype Specific and Sensitive Cell-Based Neutralization Test for Botulinum Toxin A and E.

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Bak, Nicola; Rajagopal, Shalini; Stickings, Paul; Sesardic, Dorothea

2017-07-20

Botulinum toxins (BoNTs), of which there are seven serotypes, are among the most potent neurotoxins, with serotypes A, B and E causing human botulism. Antitoxins form the first line of treatment for botulism, and functional, highly sensitive in vitro methods for toxin neutralization are needed to replace the current in vivo methods used for determination of antitoxin potency. In this preliminary proof of concept study, we report the development of a neutralization test using the neuroblastoma SiMa cell line. The assay is serotype specific for either BoNT/A or BoNT/E, which both cleave unique sequences on SNAP-25 within SiMa cells. The end point is simple immunodetection of cleaved SNAP-25 from cell lysates with antibodies detecting only the newly exposed sequence on SNAP-25. Neutralizing antibodies prevent the toxin-induced cleavage of SNAP-25. The toxin neutralization assay, with an EC50 of ~2 mIU/mL determined with a standardized reference antiserum, is more sensitive than the mouse bioassays. Relevance was demonstrated with commercial and experimental antitoxins targeting different functional domains, and of known in vivo neutralizing activities. This is the first report describing a simple, specific, in vitro cell-based assay for the detection of neutralizing antibodies against BoNT/A and BoNT/E with a sensitivity exceeding that of the mouse bioassay.

MHC Class II and Non-MHC Class II Genes Differentially Influence Humoral Immunity to Bacillus anthracis Lethal Factor and Protective Antigen

OpenAIRE

Garman, Lori; Dumas, Eric K.; Kurella, Sridevi; Hunt, Jonathan J.; Crowe, Sherry R.; Nguyen, Melissa L.; Cox, Philip M.; James, Judith A.; Farris, A. Darise

2012-01-01

Anthrax Lethal Toxin consists of Protective Antigen (PA) and Lethal Factor (LF), and current vaccination strategies focus on eliciting antibodies to PA. In human vaccination, the response to PA can vary greatly, and the response is often directed toward non-neutralizing epitopes. Variable vaccine responses have been shown to be due in part to genetic differences in individuals, with both MHC class II and other genes playing roles. Here, we investigated the relative contribution of MHC class I...

DNA aptamers as a novel approach to neutralize Staphylococcus aureus α-toxin.

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Vivekananda, Jeevalatha; Salgado, Christi; Millenbaugh, Nancy J

2014-02-14

Staphylococcus aureus is a versatile pathogen capable of causing a broad spectrum of diseases ranging from superficial skin infections to life threatening conditions such as endocarditis, septicemia, pneumonia and toxic shock syndrome. In vitro and in vivo studies identified an exotoxin, α-toxin, as a major cause of S. aureus toxicity. Because S. aureus has rapidly evolved resistance to a number of antibiotics, including methicillin, it is important to identify new therapeutic strategies, other than antibiotics, for inhibiting the harmful effects of this pathogen. Aptamers are single-stranded DNA or RNA oligonucleotides with three-dimensional folded conformations that bind with high affinity and selectivity to targets and modulate their biological functions. The goal of this study was to isolate DNA aptamers that specifically inhibit the cytotoxic activity of α-toxin. After 10 rounds of Systematic Evolution of Ligands by EXponential Enrichment (SELEX), 49 potential anti-α-toxin aptamers were identified. In vitro neutralization assays demonstrated that 4 of these 49 aptamers, AT-27, AT-33, AT-36, and AT-49, significantly inhibited α-toxin-mediated cell death in Jurkat T cells. Furthermore, RT-PCR analysis revealed that α-toxin increased the transcription of the inflammatory cytokines TNF-α and IL-17 and that anti-α-toxin aptamers AT-33 and AT-36 inhibited the upregulation of these genes. Collectively, the data suggest the feasibility of generating functionally effective aptamers against α-toxin for treatment of S. aureus infections. Published by Elsevier Inc.

Use of 51Cr release to measure the cytotoxic effects of staphylococcal leukocidin and toxin neutralization on bovine leukocytes

International Nuclear Information System (INIS)

Loeffler, D.A.; Schat, K.A.; Norcross, N.L.

1986-01-01

Leukocidin toxin from Staphylococcus aureus produces specific cytolytic effects on neutrophils and macrophages. The most commonly used method for determination of leukocidin activity is microscopic examination for characteristic morphological changes in toxin-treated cells. The 51 Cr release assay was modified to allow quantitation of the cytolytic effects of leukocidin on bovine peripheral blood neutrophils and lymphocytes. Toxin neutralization by serum and milk samples was quantitated by this method. The neutralizing abilities of the various samples were found to correlate with the levels of immunoglobulin G (IgG1) specific for leukocidin. Undiluted normal serum samples, however, were capable of partially preventing the cytotoxic effects of leukocidin. The assay was shown to be an effective means of quantitating the cytotoxic activity of leukocidin on neutrophils as well as demonstrating neutralization of cytotoxicity by milk and serum samples

Venom of the Coral Snake Micrurus clarki: Proteomic Profile, Toxicity, Immunological Cross-Neutralization, and Characterization of a Three-Finger Toxin

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Bruno Lomonte

2016-05-01

Full Text Available Micrurus clarki is an uncommon coral snake distributed from the Southeastern Pacific of Costa Rica to Western Colombia, for which no information on its venom could be found in the literature. Using a ‘venomics’ approach, proteins of at least nine families were identified, with a moderate predominance of three-finger toxins (3FTx; 48.2% over phospholipase A2 (PLA2; 36.5%. Comparison of this venom profile with those of other Micrurus species suggests that it may represent a more balanced, ‘intermediate’ type within the dichotomy between 3FTx- and PLA2-predominant venoms. M. clarki venom was strongly cross-recognized and, accordingly, efficiently neutralized by an equine therapeutic antivenom against M. nigrocinctus, revealing their high antigenic similarity. Lethal activity for mice could be reproduced by a PLA2 venom fraction, but, unexpectedly, not by fractions corresponding to 3FTxs. The most abundant venom component, hereby named clarkitoxin-I, was identified as a short-chain (type I 3FTx, devoid of lethal effect in mice, whose target remains to be defined. Its amino acid sequence of 66 residues shows high similarity with predicted sequences of venom gland transcripts described for M. fulvius, M. browni, and M. diastema.

Highly predictive support vector machine (SVM) models for anthrax toxin lethal factor (LF) inhibitors.

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Zhang, Xia; Amin, Elizabeth Ambrose

2016-01-01

Anthrax is a highly lethal, acute infectious disease caused by the rod-shaped, Gram-positive bacterium Bacillus anthracis. The anthrax toxin lethal factor (LF), a zinc metalloprotease secreted by the bacilli, plays a key role in anthrax pathogenesis and is chiefly responsible for anthrax-related toxemia and host death, partly via inactivation of mitogen-activated protein kinase kinase (MAPKK) enzymes and consequent disruption of key cellular signaling pathways. Antibiotics such as fluoroquinolones are capable of clearing the bacilli but have no effect on LF-mediated toxemia; LF itself therefore remains the preferred target for toxin inactivation. However, currently no LF inhibitor is available on the market as a therapeutic, partly due to the insufficiency of existing LF inhibitor scaffolds in terms of efficacy, selectivity, and toxicity. In the current work, we present novel support vector machine (SVM) models with high prediction accuracy that are designed to rapidly identify potential novel, structurally diverse LF inhibitor chemical matter from compound libraries. These SVM models were trained and validated using 508 compounds with published LF biological activity data and 847 inactive compounds deposited in the Pub Chem BioAssay database. One model, M1, demonstrated particularly favorable selectivity toward highly active compounds by correctly predicting 39 (95.12%) out of 41 nanomolar-level LF inhibitors, 46 (93.88%) out of 49 inactives, and 844 (99.65%) out of 847 Pub Chem inactives in external, unbiased test sets. These models are expected to facilitate the prediction of LF inhibitory activity for existing molecules, as well as identification of novel potential LF inhibitors from large datasets. Copyright © 2015 Elsevier Inc. All rights reserved.

Egg yolk antibodies for detection and neutralization of Clostridium botulinum type A neurotoxin.

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Trott, D L; Yang, M; Gonzalez, J; Larson, A E; Tepp, W H; Johnson, E A; Cook, M E

2009-05-01

The objective of this research project was to determine the usefulness of an egg antibody platform for producing materials for the detection and neutralization of botulinum type A neurotoxin. Yield estimates for detection and neutralizing antibodies produced using methods described were calculated. Antibody specific to botulinum toxoid A (aToxoid) and toxin A (aBoNT/A) was produced by immunizing hens with botulinum toxoid A (toxoid) followed by increasing amounts of botulinum neurotoxin A (BoNT/A) in Freund incomplete adjuvant. Egg yolks were extracted with polyethylene glycol (PEG) for antibody detection and neutralization experiments. A model aToxoid/toxoid immunoassay using only egg yolk antibody was developed and had a detection limit of 1 pg/ml of toxoid. In an indirect enzyme-linked immunosorbent assay of BoNT/A-specific antibody, the aBoNT/A contained more BoNT/A-specific antibody than did the aToxoid, and aBoNT/A was as effective as commercial rabbit antibody. The aToxoid provided no protection against BoNT/A in a standard mouse neutralization assay; however, 1 mg of PEG-extracted aBoNT/A neutralized 4,000 lethal doses of BoNT/A injected intraperitoneally. Based on these results, we calculated that in 1 month one hen could produce more than 100 liters of antibody detection reagents or enough antibody to neutralize approximately 11.6 million mouse lethal doses of botulinum toxin. Utilization of an egg antibody platform is potentially rapid (28 to 70 days) and scalable to kilogram quantities using current egg production facilities with as few as 1,000 hens.

Genetically modified anthrax lethal toxin safely delivers whole HIV protein antigens into the cytosol to induce T cell immunity

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Lu, Yichen; Friedman, Rachel; Kushner, Nicholas; Doling, Amy; Thomas, Lawrence; Touzjian, Neal; Starnbach, Michael; Lieberman, Judy

2000-07-01

Bacillus anthrax lethal toxin can be engineered to deliver foreign proteins to the cytosol for antigen presentation to CD8 T cells. Vaccination with modified toxins carrying 8-9 amino acid peptide epitopes induces protective immunity in mice. To evaluate whether large protein antigens can be used with this system, recombinant constructs encoding several HIV antigens up to 500 amino acids were produced. These candidate HIV vaccines are safe in animals and induce CD8 T cells in mice. Constructs encoding gag p24 and nef stimulate gag-specific CD4 proliferation and a secondary cytotoxic T lymphocyte response in HIV-infected donor peripheral blood mononuclear cells in vitro. These results lay the foundation for future clinical vaccine studies.

Binding of superantigen toxins into the CD28 homodimer interface is essential for induction of cytokine genes that mediate lethal shock.

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Gila Arad

2011-09-01

Full Text Available Bacterial superantigens, a diverse family of toxins, induce an inflammatory cytokine storm that can lead to lethal shock. CD28 is a homodimer expressed on T cells that functions as the principal costimulatory ligand in the immune response through an interaction with its B7 coligands, yet we show here that to elicit inflammatory cytokine gene expression and toxicity, superantigens must bind directly into the dimer interface of CD28. Preventing access of the superantigen to CD28 suffices to block its lethality. Mice were protected from lethal superantigen challenge by short peptide mimetics of the CD28 dimer interface and by peptides selected to compete with the superantigen for its binding site in CD28. Superantigens use a conserved β-strand/hinge/α-helix domain of hitherto unknown function to engage CD28. Mutation of this superantigen domain abolished inflammatory cytokine gene induction and lethality. Structural analysis showed that when a superantigen binds to the T cell receptor on the T cell and major histocompatibility class II molecule on the antigen-presenting cell, CD28 can be accommodated readily as third superantigen receptor in the quaternary complex, with the CD28 dimer interface oriented towards the β-strand/hinge/α-helix domain in the superantigen. Our findings identify the CD28 homodimer interface as a critical receptor target for superantigens. The novel role of CD28 as receptor for a class of microbial pathogens, the superantigen toxins, broadens the scope of pathogen recognition mechanisms.

Sub-lethal effects of Vip3A toxin on survival, development and fecundity of Heliothis virescens and Plutella xylostella.

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Gulzar, Asim; Wright, Denis J

2015-11-01

The assessment of sub-lethal effects is important to interpret the overall insecticide efficacy in controlling insect pest populations. In addition to the lethal effect, sub-lethal effects may also occur in exposed insects. Vegetative insecticidal proteins (Vips) have shown a broad spectrum of insecticidal activity against many insect pest species. In this study the sub-lethal effects of the Bacillus thuringiensis vegetative insecticidal toxin Vip3A on the development and reproduction of Heliothis virescens F. and Plutella xylostella L. were evaluated in the laboratory. The results indicated that the sub-lethal concentration of Vip3A increased the duration of the larval and pupal stages as compared with the control treatment for both species. The percent pupation and percent adult emergence were significantly lower for Vip3A-treated insects. The proportion of pairs that produced eggs and the longevity of adults were not significantly different between treatments. H. virescens and P. xylostella treated with Vip3A showed an 11 and 17 % decrease in their intrinsic rate of increase (rm) respectively compared with untreated insects. The results from this study will be helpful to develop the strategy to incorporate Vip 3A containing crops in an integrated pest management programme.

Comparative study on the ability of IgG and F(ab')2 antivenoms to neutralize lethal and myotoxic effects induced by Micrurus nigrocinctus (coral snake) venom

OpenAIRE

León Montero, Guillermo; Stiles, Bradley G.; Alape Girón, Alberto; Rojas Céspedes, Gustavo; Gutiérrez, José María

1999-01-01

A comparative study was performed on the ability of IgG and F(ab')2 antivenoms to neutralize lethal and myotoxic activities of Micrurus nigrocinctus venom. Both antivenoms were adjusted to a similar neutralizing potency in experiments where venom and antivenoms were preincubated prior to injection. No significant differences were observed between IgG and F(ab')2 antivenoms concerning neutralization of lethal effect in rescue experiments, i.e., when antivenom was administered intravenously aft...

Passive therapy with humanized anti-staphylococcal enterotoxin B antibodies attenuates systemic inflammatory response and protects from lethal pneumonia caused by staphylococcal enterotoxin B-producing Staphylococcus aureus.

Science.gov (United States)

Karau, Melissa J; Tilahun, Mulualem E; Krogman, Ashton; Osborne, Barbara A; Goldsby, Richard A; David, Chella S; Mandrekar, Jayawant N; Patel, Robin; Rajagopalan, Govindarajan

2017-10-03

Drugs such as linezolid that inhibit bacterial protein synthesis may be beneficial in treating infections caused by toxigenic Staphylococcus aureus. As protein synthesis inhibitors have no effect on preformed toxins, neutralization of pathogenic exotoxins with anti-toxin antibodies may be beneficial in conjunction with antibacterial therapy. Herein, we evaluated the efficacy of human-mouse chimeric high-affinity neutralizing anti-staphylococcal enterotoxin B (SEB) antibodies in the treatment of experimental pneumonia caused by SEB-producing S. aureus. Since HLA class II transgenic mice mount a stronger systemic immune response following challenge with SEB and are more susceptible to SEB-induced lethal toxic shock than conventional mice strains, HLA-DR3 transgenic mice were used. Lethal pneumonia caused by SEB-producing S. aureus in HLA-DR3 transgenic mice was characterized by robust T cell activation and elevated systemic levels of several pro-inflammatory cytokines and chemokines. Prophylactic administration of a single dose of linezolid 30 min prior to the onset of infection attenuated the systemic inflammatory response and protected from mortality whereas linezolid administered 60 min after the onset of infection failed to confer significant protection. Human-mouse chimeric high-affinity neutralizing anti-SEB antibodies alone, but not polyclonal human IgG, mitigated this response and protected from death when administered immediately after initiation of infection. Further, anti-SEB antibodies as well as intact polyclonal human IgG, but not its Fab or Fc fragments, protected from lethal pneumonia when followed with linezolid therapy 60 min later. In conclusion, neutralization of superantigens with high-affinity antibodies may have beneficial effects in pneumonia.

CD28: Direct and Critical Receptor for Superantigen Toxins

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Ziv Rotfogel

2013-09-01

Full Text Available Every adaptive immune response requires costimulation through the B7/CD28 axis, with CD28 on T-cells functioning as principal costimulatory receptor. Staphylococcal and streptococcal superantigen toxins hyperstimulate the T-cell-mediated immune response by orders of magnitude, inducing a lethal cytokine storm. We show that to elicit an inflammatory cytokine storm and lethality, superantigens must bind directly to CD28. Blocking access of the superantigen to its CD28 receptor with peptides mimicking the contact domains in either toxin or CD28 suffices to protect mice effectively from lethal shock. Our finding that CD28 is a direct receptor of superantigen toxins broadens the scope of microbial pathogen recognition mechanisms.

Presence of virus neutralizing antibodies in cerebral spinal fluid correlates with non-lethal rabies in dogs.

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Clement W Gnanadurai

Full Text Available Rabies is traditionally considered a uniformly fatal disease after onset of clinical manifestations. However, increasing evidence indicates that non-lethal infection as well as recovery from flaccid paralysis and encephalitis occurs in laboratory animals as well as humans.Non-lethal rabies infection in dogs experimentally infected with wild type dog rabies virus (RABV, wt DRV-Mexico correlates with the presence of high level of virus neutralizing antibodies (VNA in the cerebral spinal fluid (CSF and mild immune cell accumulation in the central nervous system (CNS. By contrast, dogs that succumbed to rabies showed only little or no VNA in the serum or in the CSF and severe inflammation in the CNS. Dogs vaccinated with a rabies vaccine showed no clinical signs of rabies and survived challenge with a lethal dose of wild-type DRV. VNA was detected in the serum, but not in the CSF of immunized dogs. Thus the presence of VNA is critical for inhibiting virus spread within the CNS and eventually clearing the virus from the CNS.Non-lethal infection with wt RABV correlates with the presence of VNA in the CNS. Therefore production of VNA within the CNS or invasion of VNA from the periphery into the CNS via compromised blood-brain barrier is important for clearing the virus infection from CNS, thereby preventing an otherwise lethal rabies virus infection.

Mouse in Vivo Neutralization of Escherichia coli Shiga Toxin 2 with Monoclonal Antibodies

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Larry H. Stanker

2013-10-01

Full Text Available Shiga toxin-producing Escherichia coli (STEC food contaminations pose serious health concerns, and have been the subject of massive food recalls. STEC has been identified as the major cause of the life-threatening complication of hemolytic uremic syndrome (HUS. Besides supportive care, there currently are no therapeutics available. The use of antibiotics for combating pathogenic E. coli is not recommended because they have been shown to stimulate toxin production. Clearing Stx2 from the circulation could potentially lessen disease severity. In this study, we tested the in vivo neutralization of Stx2 in mice using monoclonal antibodies (mAbs. We measured the biologic half-life of Stx2 in mice and determined the distribution phase or t1/2 α to be 3 min and the clearance phase or t1/2 β to be 40 min. Neutralizing mAbs were capable of clearing Stx2 completely from intoxicated mouse blood within minutes. We also examined the persistence of these mAbs over time and showed that complete protection could be passively conferred to mice 4 weeks before exposure to Stx2. The advent of better diagnositic methods and the availability of a greater arsenal of therapeutic mAbs against Stx2 would greatly enhance treatment outcomes of life threatening E. coli infections.

Tumor Targeting and Drug Delivery by Anthrax Toxin

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Christopher Bachran

2016-07-01

Full Text Available Anthrax toxin is a potent tripartite protein toxin from Bacillus anthracis. It is one of the two virulence factors and causes the disease anthrax. The receptor-binding component of the toxin, protective antigen, needs to be cleaved by furin-like proteases to be activated and to deliver the enzymatic moieties lethal factor and edema factor to the cytosol of cells. Alteration of the protease cleavage site allows the activation of the toxin selectively in response to the presence of tumor-associated proteases. This initial idea of re-targeting anthrax toxin to tumor cells was further elaborated in recent years and resulted in the design of many modifications of anthrax toxin, which resulted in successful tumor therapy in animal models. These modifications include the combination of different toxin variants that require activation by two different tumor-associated proteases for increased specificity of toxin activation. The anthrax toxin system has proved to be a versatile system for drug delivery of several enzymatic moieties into cells. This highly efficient delivery system has recently been further modified by introducing ubiquitin as a cytosolic cleavage site into lethal factor fusion proteins. This review article describes the latest developments in this field of tumor targeting and drug delivery.

Tumor Targeting and Drug Delivery by Anthrax Toxin.

Science.gov (United States)

Bachran, Christopher; Leppla, Stephen H

2016-07-01

Anthrax toxin is a potent tripartite protein toxin from Bacillus anthracis. It is one of the two virulence factors and causes the disease anthrax. The receptor-binding component of the toxin, protective antigen, needs to be cleaved by furin-like proteases to be activated and to deliver the enzymatic moieties lethal factor and edema factor to the cytosol of cells. Alteration of the protease cleavage site allows the activation of the toxin selectively in response to the presence of tumor-associated proteases. This initial idea of re-targeting anthrax toxin to tumor cells was further elaborated in recent years and resulted in the design of many modifications of anthrax toxin, which resulted in successful tumor therapy in animal models. These modifications include the combination of different toxin variants that require activation by two different tumor-associated proteases for increased specificity of toxin activation. The anthrax toxin system has proved to be a versatile system for drug delivery of several enzymatic moieties into cells. This highly efficient delivery system has recently been further modified by introducing ubiquitin as a cytosolic cleavage site into lethal factor fusion proteins. This review article describes the latest developments in this field of tumor targeting and drug delivery.

Neutralization of toxicological activities of medically-relevant Bothrops snake venoms and relevant toxins by two polyvalent bothropic antivenoms produced in Peru and Brazil.

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Estevao-Costa, Maria I; Gontijo, Silea S; Correia, Barbara L; Yarleque, Armando; Vivas-Ruiz, Dan; Rodrigues, Edith; Chávez-Olortegui, Carlos; Oliveira, Luciana S; Sanchez, Eladio F

2016-11-01

Snakebite envenoming is a neglected public pathology, affecting especially rural communities or isolated areas of tropical and subtropical Latin American countries. The parenteral administration of antivenom is the mainstay and the only validated treatment of snake bite envenoming. Here, we assess the efficacy of polyspecific anti-Bothrops serum (α-BS) produced in the Instituto Nacional de Salud (INS, Peru) and at the Fundação Ezequiel Dias (FUNED, Brazil), to neutralize the main toxic activities induced by five medically-relevant venoms of: Bothrops atrox, B. barnetti, and B. pictus from Peru, and the Brazilian B. jararaca and B. leucurus, all of them inhabiting different geographical locations. Protein electrophoretic patterns of these venoms showed significant differences in composition, number and intensity of bands. Another goal was to evaluate the efficacy and safety of lyophilized α-BS developed at INS to neutralize the detrimental effects of these venoms using in vivo and in vitro assays. The availability of lyophilized α-BS has relevant significance in its distribution to distant rural communities where the access to antivenom in health facilities is more difficult. Despite the fact that different antigen mixtures were used for immunization during antivenom production, our data showed high toxin-neutralizing activity of α-BS raised against Bothrops venoms. Moreover, the antivenom cross-reacted even against venoms not included in the immunization mixture. Furthermore, we have evaluated the efficacy of both α-BS to neutralize key toxic compounds belonging to the predominant protein families of Bothrops snakes. Most significantly, both α-BS cross-specifically neutralized the main toxicological activities e.g. lethality and hemorrhage induced by these venoms. Thus, our data indicate that both α-BS are equally effective to treat snake bite victims inflicted by Bothrops snakes particularly B. atrox, responsible for the largest numbers of human

Development of a recombinant toxin fragment vaccine for Clostridium difficile infection.

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Karczewski, Jerzy; Zorman, Julie; Wang, Su; Miezeiewski, Matthew; Xie, Jinfu; Soring, Keri; Petrescu, Ioan; Rogers, Irene; Thiriot, David S; Cook, James C; Chamberlin, Mihaela; Xoconostle, Rachel F; Nahas, Debbie D; Joyce, Joseph G; Bodmer, Jean-Luc; Heinrichs, Jon H; Secore, Susan

2014-05-19

Clostridium difficile infection (CDI) is the major cause of antibiotic-associated diarrhea and pseudomembranous colitis, a disease associated with significant morbidity and mortality. The disease is mostly of nosocomial origin, with elderly patients undergoing anti-microbial therapy being particularly at risk. C. difficile produces two large toxins: Toxin A (TcdA) and Toxin B (TcdB). The two toxins act synergistically to damage and impair the colonic epithelium, and are primarily responsible for the pathogenesis associated with CDI. The feasibility of toxin-based vaccination against C. difficile is being vigorously investigated. A vaccine based on formaldehyde-inactivated Toxin A and Toxin B (toxoids) was reported to be safe and immunogenic in healthy volunteers and is now undergoing evaluation in clinical efficacy trials. In order to eliminate cytotoxic effects, a chemical inactivation step must be included in the manufacturing process of this toxin-based vaccine. In addition, the large-scale production of highly toxic antigens could be a challenging and costly process. Vaccines based on non-toxic fragments of genetically engineered versions of the toxins alleviate most of these limitations. We have evaluated a vaccine assembled from two recombinant fragments of TcdB and explored their potential as components of a novel experimental vaccine against CDI. Golden Syrian hamsters vaccinated with recombinant fragments of TcdB combined with full length TcdA (Toxoid A) developed high titer IgG responses and potent neutralizing antibody titers. We also show here that the recombinant vaccine protected animals against lethal challenge with C. difficile spores, with efficacy equivalent to the toxoid vaccine. The development of a two-segment recombinant vaccine could provide several advantages over toxoid TcdA/TcdB such as improvements in manufacturability. Copyright © 2014 Elsevier Ltd. All rights reserved.

Tumor Targeting and Drug Delivery by Anthrax Toxin

OpenAIRE

Bachran, Christopher; Leppla, Stephen H.

2016-01-01

Anthrax toxin is a potent tripartite protein toxin from Bacillus anthracis. It is one of the two virulence factors and causes the disease anthrax. The receptor-binding component of the toxin, protective antigen, needs to be cleaved by furin-like proteases to be activated and to deliver the enzymatic moieties lethal factor and edema factor to the cytosol of cells. Alteration of the protease cleavage site allows the activation of the toxin selectively in response to the presence of tumor-associ...

Clostridium difficile chimeric toxin receptor binding domain vaccine induced protection against different strains in active and passive challenge models.

Science.gov (United States)

Tian, Jing-Hui; Glenn, Gregory; Flyer, David; Zhou, Bin; Liu, Ye; Sullivan, Eddie; Wu, Hua; Cummings, James F; Elllingsworth, Larry; Smith, Gale

2017-07-24

Clostridium difficile is the number one cause of nosocomial antibiotic-associated diarrhea in developed countries. Historically, pathogenesis was attributed two homologous glucosylating toxins, toxin-A (TcdA) and toxin-B (TcdB). Over the past decade, however, highly virulent epidemic strains of C. difficile (B1/NAP1/027) have emerged and are linked to an increase in morbidity and mortality. Increased virulence is attributed to multiple factors including: increased production of A- and B-toxins; production of binary toxin (CDT); and the emergence of more toxic TcdB variants (TcdB (027) ). TcdB (027) is more cytotoxicity to cells; causes greater tissue damage and toxicity in animals; and is antigenically distinct from historical TcdB (TcdB (003) ). Broadly protective vaccines and therapeutic antibody strategies, therefore, may target TcdA, TcdB variants and CDT. To facilitate the generation of multivalent toxin-based C. difficile vaccines and therapeutic antibodies, we have generated fusion proteins constructed from the receptor binding domains (RBD) of TcdA, TcdB (003) , TcdB (027) and CDT. Herein, we describe the development of a trivalent toxin (T-toxin) vaccine (CDTb/TcdB (003) /TcdA) and quadravalent toxin (Q-toxin) vaccine (CDTb/TcB (003) /TcdA/TcdB (027) ) fusion proteins that retain the protective toxin neutralizing epitopes. Active immunization of mice or hamsters with T-toxin or Q-toxin fusion protein vaccines elicited the generation of toxin neutralizing antibodies to each of the toxins. Hamsters immunized with the Q-toxin vaccine were broadly protected against spore challenge with historical C. difficile 630 (toxinotype 0/ribotype 003) and epidemic NAP1 (toxinotype III/ribotype 027) strains. Fully human polyclonal antitoxin IgG was produced by immunization of transgenic bovine with these fusion proteins. In passive transfer studies, mice were protected against lethal toxin challenge. Hamsters treated with human antitoxin IgG were completely protected when

Potent antitumor activity of a urokinase-activated engineered anthrax toxin

Science.gov (United States)

Liu, Shihui; Aaronson, Hannah; Mitola, David J.; Leppla, Stephen H.; Bugge, Thomas H.

2003-01-01

The acquisition of cell-surface urokinase plasminogen activator activity is a hallmark of malignancy. We generated an engineered anthrax toxin that is activated by cell-surface urokinase in vivo and displays limited toxicity to normal tissue but broad and potent tumoricidal activity. Native anthrax toxin protective antigen, when administered with a chimeric anthrax toxin lethal factor, Pseudomonas exotoxin fusion protein, was extremely toxic to mice, causing rapid and fatal organ damage. Replacing the furin activation sequence in anthrax toxin protective antigen with an artificial peptide sequence efficiently activated by urokinase greatly attenuated toxicity to mice. In addition, the mutation conferred cell-surface urokinase-dependent toxin activation in vivo, as determined by using a panel of plasminogen, plasminogen activator, plasminogen activator receptor, and plasminogen activator inhibitor-deficient mice. Surprisingly, toxin activation critically depended on both urokinase plasminogen activator receptor and plasminogen in vivo, showing that both proteins are essential cofactors for the generation of cell-surface urokinase. The engineered toxin displayed potent tumor cell cytotoxicity to a spectrum of transplanted tumors of diverse origin and could eradicate established solid tumors. This tumoricidal activity depended strictly on tumor cell-surface plasminogen activation. The data show that a simple change of protease activation specificity converts anthrax toxin from a highly lethal to a potent tumoricidal agent.

Delayed Toxicity Associated with Soluble Anthrax Toxin Receptor Decoy-Ig Fusion Protein Treatment

Science.gov (United States)

Cote, Christopher; Welkos, Susan; Manchester, Marianne; Young, John A. T.

2012-01-01

Soluble receptor decoy inhibitors, including receptor-immunogloubulin (Ig) fusion proteins, have shown promise as candidate anthrax toxin therapeutics. These agents act by binding to the receptor-interaction site on the protective antigen (PA) toxin subunit, thereby blocking toxin binding to cell surface receptors. Here we have made the surprising observation that co-administration of receptor decoy-Ig fusion proteins significantly delayed, but did not protect, rats challenged with anthrax lethal toxin. The delayed toxicity was associated with the in vivo assembly of a long-lived complex comprised of anthrax lethal toxin and the receptor decoy-Ig inhibitor. Intoxication in this system presumably results from the slow dissociation of the toxin complex from the inhibitor following their prolonged circulation. We conclude that while receptor decoy-Ig proteins represent promising candidates for the early treatment of B. anthracis infection, they may not be suitable for therapeutic use at later stages when fatal levels of toxin have already accumulated in the bloodstream. PMID:22511955

Stabilization of a recombinant ricin toxin A subunit vaccine through lyophilization.

Science.gov (United States)

Hassett, Kimberly J; Cousins, Megan C; Rabia, Lilia A; Chadwick, Chrystal M; O'Hara, Joanne M; Nandi, Pradyot; Brey, Robert N; Mantis, Nicholas J; Carpenter, John F; Randolph, Theodore W

2013-10-01

Lyophilization was used to prepare dry, glassy solid vaccine formulations of recombinant ricin toxin A-chain containing suspensions of colloidal aluminum hydroxide adjuvant. Four lyophilized formulations were prepared by using combinations of rapid or slow cooling during lyophilization and one of two buffers, histidine or ammonium acetate. Trehalose was used as the stabilizing excipient. Aggregation of the colloidal aluminum hydroxide suspension was reduced in formulations processed with a rapid cooling rate. Aluminum hydroxide particle size distributions, glass transition temperatures, water contents, and immunogenicities of lyophilized vaccines were independent of incubation time at 40 °C for up to 15 weeks. Mice immunized with reconstituted ricin toxin subunit A (RTA) vaccines produced RTA-specific antibodies and toxin-neutralizing antibodies (TNAs) regardless of the length of high temperature vaccine storage or the degree of aluminum adjuvant aggregation that occurred during lyophilization. In murine studies, lyophilized formulations of vaccines conferred protection against exposure to lethal doses of ricin, even after the lyophilized formulations had been stored at 40 °C for 4 weeks. A corresponding liquid formulation of vaccine stored at 40 °C elicited RTA-specific antibody titers but failed to confer immunity during a ricin challenge. Copyright © 2013 Elsevier B.V. All rights reserved.

Radiolabelling of cholera toxin

International Nuclear Information System (INIS)

Santos, R.G.; Neves, Nicoli M.J.; Abdalla, L.F.; Brandao, R.L.; Etchehebehere, L.; Lima, M.E. de; Nicoli, J.R.

1999-01-01

Binding of cholera toxin to ganglioside receptors of enterocyte microvilli catalyzes the activation of adenylate cyclase causing a rise in cAMP which final result is a copious diarrhea. Saccharomyces boulardii, a nonpathogenic yeast has been used to prevent diarrhea. Although the antidiarrheic properties of S. boulardii are widely recognized, this yeast has been used on empirical basis, and the mechanism of this protective effect is unknown. The addition of cholera toxin to S. boulardii induces the raising of cAMP that triggers the activation of neutral trehalase. This suggests that toxin specifically binding to cells, is internalized and active the protein phosphorylation cascade. Our objective is labeling the cholera toxin to verify the presence of binding sites on yeast cell surfaces for the cholera toxin. Cholera toxin was radiolabelled with Na 125 I by a chloramine-T method modified from Cuatrecasas and Griffiths et alii. The 125 I-Cholera toxin showed a specific radioactivity at about 1000 cpm/fmol toxin. Biological activity of labeled cholera toxin measured by trehalase activation was similar to the native toxin. (author)

Toxin-Based Therapeutic Approaches

Science.gov (United States)

Shapira, Assaf; Benhar, Itai

2010-01-01

Protein toxins confer a defense against predation/grazing or a superior pathogenic competence upon the producing organism. Such toxins have been perfected through evolution in poisonous animals/plants and pathogenic bacteria. Over the past five decades, a lot of effort has been invested in studying their mechanism of action, the way they contribute to pathogenicity and in the development of antidotes that neutralize their action. In parallel, many research groups turned to explore the pharmaceutical potential of such toxins when they are used to efficiently impair essential cellular processes and/or damage the integrity of their target cells. The following review summarizes major advances in the field of toxin based therapeutics and offers a comprehensive description of the mode of action of each applied toxin. PMID:22069564

Chimeric anti-staphylococcal enterotoxin B antibodies and lovastatin act synergistically to provide in vivo protection against lethal doses of SEB.

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Mulualem E Tilahun

Full Text Available Staphylococcal enterotoxin B (SEB is one of a family of toxins secreted by Staphylococcus aureus that act as superantigens, activating a large fraction of the T-cell population and inducing production of high levels of inflammatory cytokines that can cause toxic shock syndrome (TSS and death. Extracellular engagement of the TCR of T-cells and class II MHC of antigen presenting cells by SEB triggers the activation of many intracellular signaling processes. We engineered chimeric antibodies to block the extracellular engagement of cellular receptors by SEB and used a statin to inhibit intracellular signaling. Chimeric human-mouse antibodies directed against different neutralizing epitopes of SEB synergistically inhibited its activation of human T-cells in vitro. In the in vivo model of lethal toxic shock syndrome (TSS in HLA-DR3 transgenic mice, two of these antibodies conferred significant partial protection when administered individually, but offered complete protection in a synergistic manner when given together. Similarly, in vivo, lovastatin alone conferred only partial protection from TSS similar to single anti-SEB antibodies. However, used in combination with one chimeric neutralizing anti-SEB antibody, lovastatin provided complete protection against lethal TSS in HLA-DR3 transgenic mice. These experiments demonstrate that in vivo protection against lethal doses of SEB can be achieved by a statin of proven clinical safety and chimeric human-mouse antibodies, agents now widely used and known to be of low immunogenicity in human hosts.

Novel Clostridium difficile Anti-Toxin (TcdA and TcdB Humanized Monoclonal Antibodies Demonstrate In Vitro Neutralization across a Broad Spectrum of Clinical Strains and In Vivo Potency in a Hamster Spore Challenge Model.

Directory of Open Access Journals (Sweden)

Hongyu Qiu

Full Text Available Clostridium difficile (C. difficile infection (CDI is the main cause of nosocomial antibiotic-associated colitis and increased incidence of community-associated diarrhea in industrialized countries. At present, the primary treatment of CDI is antibiotic administration, which is effective but often associated with recurrence, especially in the elderly. Pathogenic strains produce enterotoxin, toxin A (TcdA, and cytotoxin, toxin B (TcdB, which are necessary for C. difficile induced diarrhea and gut pathological changes. Administration of anti-toxin antibodies provides an alternative approach to treat CDI, and has shown promising results in preclinical and clinical studies. In the current study, several humanized anti-TcdA and anti-TcdB monoclonal antibodies were generated and their protective potency was characterized in a hamster infection model. The humanized anti-TcdA (CANmAbA4 and anti-TcdB (CANmAbB4 and CANmAbB1 antibodies showed broad spectrum in vitro neutralization of toxins from clinical strains and neutralization in a mouse toxin challenge model. Moreover, co-administration of humanized antibodies (CANmAbA4 and CANmAbB4 cocktail provided a high level of protection in a dose dependent manner (85% versus 57% survival at day 22 for 50 mg/kg and 20 mg/kg doses, respectively in a hamster gastrointestinal infection (GI model. This study describes the protective effects conferred by novel neutralizing anti-toxin monoclonal antibodies against C. difficile toxins and their potential as therapeutic agents in treating CDI.

Botulinum toxin: bioweapon & magic drug.

Science.gov (United States)

Dhaked, Ram Kumar; Singh, Manglesh Kumar; Singh, Padma; Gupta, Pallavi

2010-11-01

Botulinum neurotoxins, causative agents of botulism in humans, are produced by Clostridium botulinum, an anaerobic spore-former Gram positive bacillus. Botulinum neurotoxin poses a major bioweapon threat because of its extreme potency and lethality; its ease of production, transport, and misuse; and the need for prolonged intensive care among affected persons. A single gram of crystalline toxin, evenly dispersed and inhaled, can kill more than one million people. The basis of the phenomenal potency of botulinum toxin is enzymatic; the toxin is a zinc proteinase that cleaves neuronal vesicle associated proteins responsible for acetylcholine release into the neuromuscular junction. As a military or terrorist weapon, botulinum toxin could be disseminated via aerosol or by contamination of water or food supplies, causing widespread casualties. A fascinating aspect of botulinum toxin research in recent years has been development of the most potent toxin into a molecule of significant therapeutic utility . It is the first biological toxin which is licensed for treatment of human diseases. In the late 1980s, Canada approved use of the toxin to treat strabismus, in 2001 in the removal of facial wrinkles and in 2002, the FDA in the United States followed suit. The present review focuses on both warfare potential and medical uses of botulinum neurotoxin.

Clinical Study of New Tetravalent (Type A, B, E, and F) Botulinum Toxoid Vaccine Derived from M Toxin in Japan.

Science.gov (United States)

Torii, Yasushi; Sugimoto, Nakaba; Kohda, Tomoko; Kozaki, Shunji; Morokuma, Kazunori; Horikawa, Yoshikane; Ginnaga, Akihiro; Yamamoto, Akihiko; Takahashi, Motohide

2017-07-24

Botulinum toxin is the most poisonous substance known, and is believed to be a highly lethal as a biological weapon; researchers of the toxin are exposed to this hazard. Botulinum toxoid vaccines have been produced and used in Japan. However, since clinical studies involving these vaccines were conducted before establishment of the Ethical Guidelines for Clinical Research in Japan, their immunogenicity and safety were not systematically assessed. In this study, we produced a new tetravalent (type A, B, E, and F) botulinum toxoid vaccine, the first ever to be derived from M toxin, and conducted quality control tests with reference to the Minimum Requirements in Japan for adsorbed tetanus toxoid vaccine. Subsequently, a clinical study using the new vaccine in 48 healthy adult volunteers was conducted according to the guidelines in Japan. No clinically serious adverse event was noted. Neutralizing antibody titers for each type of toxin in the participants' sera, 1 month after the 4th injection were more than 0.25 IU/mL, indicating sufficient protection. This study demonstrated that the vaccine has marked immunogenicity and is safe for use in humans.

A STUDY OF IMMUNOGENIC AND PROTECTIVE PROPERTIES OF THE HEAT-STABLE LETHAL TOXIN OF YERSINIA PSEUDOTUBERCULOSIS AND ITS EFFECTS UPON HEMATOLOGICAL AND BLOOD CYTOKINE PARAMETERS OF LABORATORY MICE

Directory of Open Access Journals (Sweden)

A. V. Tsybulsky

2014-01-01

Full Text Available The article presents some data concerning antigenic and immunogenic properties of the lethal heat-stable toxin (HST from Yersinia pseudotuberculosis, a protein with molecular weight of 45 kDa. The mice,following double immunization with HST at a dose of 0.1 mg per mouse, displayed higher antibody production, in comparison with a dose of 0.01 mg/mouse. The appropriate differences were revealed with regard ofleukocyte responses, i.e., development of leukopenia, neutropenia, lymphopenia upon immunization with the 0.01 mg of HST per mouse, whereas leukocytosis, and increase in lymphocytes and monocytes was detected after a dose of 0.1 mg/mouse. We detected some doseependent differences in cytokine-modulating activity. I.e., at HST dose of 0.01 mg per mouse, we detected mostly proinflammatory, acutehase responses, whereas a dose of 0.1 mg/mice caused induction of . IFNγ and cytokines promoting lymphocyte proliferation and antibody production by day +17. Upon double immunization of mice, the toxin showed protective properties when injecting them with lethal dose of Y. pseudotuberculosis. A lagging activation of antibody producers duringHST response suggests a need for searching effective adjuvant tools of enhancement and acceleration of specific humoral immune reactions against this antigen.

Toxin-Based Therapeutic Approaches

Directory of Open Access Journals (Sweden)

Itai Benhar

2010-10-01

Full Text Available Protein toxins confer a defense against predation/grazing or a superior pathogenic competence upon the producing organism. Such toxins have been perfected through evolution in poisonous animals/plants and pathogenic bacteria. Over the past five decades, a lot of effort has been invested in studying their mechanism of action, the way they contribute to pathogenicity and in the development of antidotes that neutralize their action. In parallel, many research groups turned to explore the pharmaceutical potential of such toxins when they are used to efficiently impair essential cellular processes and/or damage the integrity of their target cells. The following review summarizes major advances in the field of toxin based therapeutics and offers a comprehensive description of the mode of action of each applied toxin.

Enhanced neutralization potency of botulinum neurotoxin antibodies using a red blood cell-targeting fusion protein.

Directory of Open Access Journals (Sweden)

Sharad P Adekar

2011-03-01

Full Text Available Botulinum neurotoxin (BoNT potently inhibits cholinergic signaling at the neuromuscular junction. The ideal countermeasures for BoNT exposure are monoclonal antibodies or BoNT antisera, which form BoNT-containing immune complexes that are rapidly cleared from the general circulation. Clearance of opsonized toxins may involve complement receptor-mediated immunoadherence to red blood cells (RBC in primates or to platelets in rodents. Methods of enhancing immunoadherence of BoNT-specific antibodies may increase their potency in vivo. We designed a novel fusion protein (FP to link biotinylated molecules to glycophorin A (GPA on the RBC surface. The FP consists of an scFv specific for murine GPA fused to streptavidin. FP:mAb:BoNT complexes bound specifically to the RBC surface in vitro. In a mouse model of BoNT neutralization, the FP increased the potency of single and double antibody combinations in BoNT neutralization. A combination of two antibodies with the FP gave complete neutralization of 5,000 LD50 BoNT in mice. Neutralization in vivo was dependent on biotinylation of both antibodies and correlated with a reduction of plasma BoNT levels. In a post-exposure model of intoxication, FP:mAb complexes gave complete protection from a lethal BoNT/A1 dose when administered within 2 hours of toxin exposure. In a pre-exposure prophylaxis model, mice were fully protected for 72 hours following administration of the FP:mAb complex. These results demonstrate that RBC-targeted immunoadherence through the FP is a potent enhancer of BoNT neutralization by antibodies in vivo.

Radiolabelling of cholera toxin

Energy Technology Data Exchange (ETDEWEB)

Santos, R.G.; Neves, Nicoli M.J. [Centro de Desenvolvimento da Tecnologia Nuclear (CDTN), Belo Horizonte, MG (Brazil); Abdalla, L.F.; Brandao, R.L.; Etchehebehere, L. [Ouro Preto Univ., MG (Brazil). Escola de Farmacia. Lab. de Fisiologia e Bioquimica de Microorganismos; Lima, M.E. de [Minas Gerais Univ., Belo Horizonte, MG (Brazil). Inst. de Ciencias Biologicas. Dept. de Bioquimica e Imunologia; Nicoli, J.R. [Minas Gerais Univ., Belo Horizonte, MG (Brazil). Inst. de Ciencias Biologicas. Dept. de Microbiologia

1999-11-01

Binding of cholera toxin to ganglioside receptors of enterocyte microvilli catalyzes the activation of adenylate cyclase causing a rise in cAMP which final result is a copious diarrhea. Saccharomyces boulardii, a nonpathogenic yeast has been used to prevent diarrhea. Although the antidiarrheic properties of S. boulardii are widely recognized, this yeast has been used on empirical basis, and the mechanism of this protective effect is unknown. The addition of cholera toxin to S. boulardii induces the raising of cAMP that triggers the activation of neutral trehalase. This suggests that toxin specifically binding to cells, is internalized and active the protein phosphorylation cascade. Our objective is labeling the cholera toxin to verify the presence of binding sites on yeast cell surfaces for the cholera toxin. Cholera toxin was radiolabelled with Na {sup 125} I by a chloramine-T method modified from Cuatrecasas and Griffiths et alii. The {sup 125} I-Cholera toxin showed a specific radioactivity at about 1000 cpm/fmol toxin. Biological activity of labeled cholera toxin measured by trehalase activation was similar to the native toxin. (author) 5 refs., 3 figs.; e-mail: nevesmj at urano.cdtn.br

Anthrax lethal toxin disrupts intestinal barrier function and causes systemic infections with enteric bacteria.

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Chen Sun

Full Text Available A variety of intestinal pathogens have virulence factors that target mitogen activated protein kinase (MAPK signaling pathways, including Bacillus anthracis. Anthrax lethal toxin (LT has specific proteolytic activity against the upstream regulators of MAPKs, the MAPK kinases (MKKs. Using a murine model of intoxication, we show that LT causes the dose-dependent disruption of intestinal epithelial integrity, characterized by mucosal erosion, ulceration, and bleeding. This pathology correlates with an LT-dependent blockade of intestinal crypt cell proliferation, accompanied by marked apoptosis in the villus tips. C57BL/6J mice treated with intravenous LT nearly uniformly develop systemic infections with commensal enteric organisms within 72 hours of administration. LT-dependent intestinal pathology depends upon its proteolytic activity and is partially attenuated by co-administration of broad spectrum antibiotics, indicating that it is both a cause and an effect of infection. These findings indicate that targeting of MAPK signaling pathways by anthrax LT compromises the structural integrity of the mucosal layer, serving to undermine the effectiveness of the intestinal barrier. Combined with the well-described immunosuppressive effects of LT, this disruption of the intestinal barrier provides a potential mechanism for host invasion via the enteric route, a common portal of entry during the natural infection cycle of Bacillus anthracis.

Toxins not neutralized by brown snake antivenom

International Nuclear Information System (INIS)

Judge, Roopwant K.; Henry, Peter J.; Mirtschin, Peter; Jelinek, George; Wilce, Jacqueline A.

2006-01-01

The Australian snakes of the genus Pseudonaja (dugite, gwardar and common brown) account for the majority of snake bite related deaths in Australia. Without antivenom treatment, the risk of mortality is significant. There is an accumulating body of evidence to suggest that the efficacy of the antivenom is limited. The current study investigates the protein constituents recognized by the antivenom using 2-DE, immuno-blot techniques and rat tracheal organ bath assays. The 2-DE profiles for all three snake venoms were similar, with major species visualized at 78-132 kDa, 32-45 kDa and 6-15 kDa. Proteins characterized by LC-MS/MS revealed a coagulant toxin (∼42 kDa) and coagulant peptide (∼6 kDa), as well as two PLA 2 (∼14 kDa). Peptides isolated from ∼78 kDa and 15-32 kDa protein components showed no similarity to known protein sequences. Protein recognition by the antivenom occurred predominantly for the higher molecular weight components with little recognition of 6-32 kDa MW species. The ability of antivenom to neutralize venom activity was also investigated using rat tracheal organ bath assays. The venoms of Pseudonaja affinis affinis and Pseudonaja nuchalis incited a sustained, significant contraction of the trachea. These contractions were attributed to PLA 2 enzymatic activity as pre-treatment with the PLA 2 inhibitor 4-BPB attenuated the venom-induced contractions. The venom of Pseudonaja textilis incited tracheal contractility through a non-PLA 2 enzymatic activity. Neither activity was attenuated by the antivenom treatment. These results represent the first proteomic investigation of the venoms from the snakes of the genus Pseudonaja, revealing a possible limitation of the brown snake antivenom in binding to the low MW protein components

Toxin-Based Therapeutic Approaches

OpenAIRE

Itai Benhar; Assaf Shapira

2010-01-01

Protein toxins confer a defense against predation/grazing or a superior pathogenic competence upon the producing organism. Such toxins have been perfected through evolution in poisonous animals/plants and pathogenic bacteria. Over the past five decades, a lot of effort has been invested in studying their mechanism of action, the way they contribute to pathogenicity and in the development of antidotes that neutralize their action. In parallel, many research groups turned to explore the pharmac...

Tumor therapy with a urokinase plasminogen activator-activated anthrax lethal toxin alone and in combination with paclitaxel.

Science.gov (United States)

Wein, Alexander N; Liu, Shihui; Zhang, Yi; McKenzie, Andrew T; Leppla, Stephen H

2013-02-01

PA-U2, an engineered anthrax protective antigen that is activated by urokinase was combined with wildtype lethal factor in the treatment of Colo205 colon adenocarcinoma in vitro and B16-BL6 mouse melanoma in vitro and in vivo. This therapy was also tested in combination with the small molecule paclitaxel, based on prior reports suggesting synergy between ERK1/2 inhibition and chemotherapeutics. Colo205 was sensitive to PA-U2/LF while B16-BL6 was not. For the combination treatment of B16-BL6, paclitaxel showed a dose response in vitro, but cells remained resistant to PA-U2/LF even in the presence of paclitaxel. In vivo, each therapy slowed tumor progression, and an additive effect between the two was observed. Since LF targets tumor vasculature while paclitaxel is an antimitotic, it is possible the agents were acting against different cells in the stroma, precluding a synergistic effect. The engineered anthrax toxin PA-U2/LF warrants further development and testing, possibly in combination with an antiangiogenesis therapy such as sunitinib or sorafinib.

Why do we study animal toxins?

Science.gov (United States)

ZHANG, Yun

2015-01-01

Venom (toxins) is an important trait evolved along the evolutionary tree of animals. Our knowledges on venoms, such as their origins and loss, the biological relevance and the coevolutionary patterns with other organisms are greatly helpful in understanding many fundamental biological questions, i.e., the environmental adaptation and survival competition, the evolution shaped development and balance of venoms, and the sophisticated correlations among venom, immunity, body power, intelligence, their genetic basis, inherent association, as well as the cost-benefit and trade-offs of biological economy. Lethal animal envenomation can be found worldwide. However, from foe to friend, toxin studies have led lots of important discoveries and exciting avenues in deciphering and fighting human diseases, including the works awarded the Nobel Prize and lots of key clinic therapeutics. According to our survey, so far, only less than 0.1% of the toxins of the venomous animals in China have been explored. We emphasize on the similarities shared by venom and immune systems, as well as the studies of toxin knowledge-based physiological toxin-like proteins/peptides (TLPs). We propose the natural pairing hypothesis. Evolution links toxins with humans. Our mission is to find out the right natural pairings and interactions of our body elements with toxins, and with endogenous toxin-like molecules. Although, in nature, toxins may endanger human lives, but from a philosophical point of view, knowing them well is an effective way to better understand ourselves. So, this is why we study toxins. PMID:26228472

Therapeutic Approaches of Botulinum Toxin in Gynecology

OpenAIRE

Marius Alexandru Moga; Oana Gabriela Dimienescu; Andreea Bălan; Ioan Scârneciu; Barna Barabaș; Liana Pleș

2018-01-01

Botulinum toxins (BoNTs) are produced by several anaerobic species of the genus Clostridium and, although they were originally considered lethal toxins, today they find their usefulness in the treatment of a wide range of pathologies in various medical specialties. Botulinum neurotoxin has been identified in seven different isoforms (BoNT-A, BoNT-B, BoNT-C, BoNT-D, BoNT-E, BoNT-F, and BoNT-G). Neurotoxigenic Clostridia can produce more than 40 different BoNT subtypes and, recently, a new BoNT...

A simple electroelution method for rapid protein purification: isolation and antibody production of alpha toxin from Clostridium septicum

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Lorena Vázquez-Iglesias

2017-06-01

Full Text Available Clostridium septicum produces a number of diseases in human and farm animals which, in most of the cases, are fatal without clinical intervention. Alpha toxin is an important agent and the unique lethal virulent factor produced by Clostridium septicum. This toxin is haemolytic, highly lethal and necrotizing activities but is being used as an antigen to develop animal vaccines. The aim of this study was to isolate the alpha toxin of Clostridium septicum and produce highly specific antibodies against it. In this work, we have developed a simple and efficient method for alpha toxin purification, based on electroelution that can be used as a time-saving method for purifying proteins. This technique avoids contamination by other proteins that could appear during other protein purification techniques such chromatography. The highly purified toxin was used to produce polyclonal antibodies. The specificity of the antibodies was tested by western blot and these antibodies can be applied to the quantitative determination of alpha toxin by slot blot.

In Vivo Neutralization of α-Cobratoxin with High-Affinity Llama Single-Domain Antibodies (VHHs) and a VHH-Fc Antibody

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Richard, Gabrielle; Meyers, Ashley J.; McLean, Michael D.; Arbabi-Ghahroudi, Mehdi; MacKenzie, Roger; Hall, J. Christopher

2013-01-01

Small recombinant antibody fragments (e.g. scFvs and VHHs), which are highly tissue permeable, are being investigated for antivenom production as conventional antivenoms consisting of IgG or F(ab’)2 antibody fragments do not effectively neutralize venom toxins located in deep tissues. However, antivenoms composed entirely of small antibody fragments may have poor therapeutic efficacy due to their short serum half-lives. To increase serum persistence and maintain tissue penetration, we prepared low and high molecular mass antivenom antibodies. Four llama VHHs were isolated from an immune VHH-displayed phage library and were shown to have high affinity, in the low nM range, for α-cobratoxin (α–Cbtx), the most lethal component of Naja kaouthia venom. Subsequently, our highest affinity VHH (C2) was fused to a human Fc fragment to create a VHH2-Fc antibody that would offer prolonged serum persistence. After in planta (Nicotiana benthamiana) expression and purification, we show that our VHH2-Fc antibody retained high affinity binding to α–Cbtx. Mouse α–Cbtx challenge studies showed that our highest affinity VHHs (C2 and C20) and the VHH2-Fc antibody effectively neutralized lethality induced by α–Cbtx at an antibody:toxin molar ratio as low as ca. 0.75×:1. Further research towards the development of an antivenom therapeutic involving these anti-α-Cbtx VHHs and VHH2-Fc antibody molecules should involve testing them as a combination, to determine whether they maintain tissue penetration capability and low immunogenicity, and whether they exhibit improved serum persistence and therapeutic efficacy. PMID:23894495

Integrative characterization of the venom of the coral snake Micrurus dumerilii (Elapidae) from Colombia: Proteome, toxicity, and cross-neutralization by antivenom.

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Rey-Suárez, Paola; Núñez, Vitelbina; Fernández, Julián; Lomonte, Bruno

2016-03-16

In Colombia, nearly 2.8% of the 4200 snakebite accidents recorded annually are inflicted by coral snakes (genus Micrurus). Micrurus dumerilii has a broad distribution in this country, especially in densely populated areas. The proteomic profile of its venom was here studied by a bottom-up approach combining RP-HPLC, SDS-PAGE and MALDI-TOF/TOF. Venom proteins were assigned to eleven families, the most abundant being phospholipases A2 (PLA2; 52.0%) and three-finger toxins (3FTx; 28.1%). This compositional profile shows that M. dumerilii venom belongs to the 'PLA2-rich' phenotype, in the recently proposed dichotomy for Micrurus venoms. Enzymatic and toxic venom activities correlated with protein family abundances. Whole venom induced a conspicuous myotoxic, cytotoxic and anticoagulant effect, and was mildly edematogenic and proteolytic, whereas it lacked hemorrhagic activity. Some 3FTxs and PLA2s reproduced the lethal effect of venom. A coral snake antivenom to Micrurus nigrocinctus demonstrated significant cross-recognition of M. dumerilii venom proteins, and accordingly, ability to neutralize its lethal effect. The combined compositional, functional, and immunological data here reported for M. dumerilii venom may contribute to a better understanding of these envenomings, and support the possible use of anti-M. nigrocinctus coral snake antivenom in their treatment. Coral snakes represent a highly diversified group of elapids in the New World, with nearly 70 species within the genus Micrurus. Owing to their scarce yields, the biochemical composition and toxic activities of coral snake venoms have been less well characterized than those of viperid species. In this work, an integrative view of the venom of M. dumerilii, a medically relevant coral snake from Colombia, was obtained by a combined proteomic, functional, and immunological approach. The venom contains proteins from at least eleven families, with a predominance of phospholipases A2 (PLA2), followed by three

Role of the Antigen Capture Pathway in the Induction of a Neutralizing Antibody Response to Anthrax Protective Antigen

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Anita Verma

2018-02-01

Full Text Available Toxin neutralizing antibodies represent the major mode of protective immunity against a number of toxin-mediated bacterial diseases, including anthrax; however, the cellular mechanisms that lead to optimal neutralizing antibody responses remain ill defined. Here we show that the cellular binding pathway of anthrax protective antigen (PA, the binding component of anthrax toxin, determines the toxin neutralizing antibody response to this antigen. PA, which binds cellular receptors and efficiently enters antigen-presenting cells by receptor-mediated endocytosis, was found to elicit robust anti-PA IgG and toxin neutralizing antibody responses. In contrast, a receptor binding-deficient mutant of PA, which does not bind receptors and only inefficiently enters antigen-presenting cells by macropinocytosis, elicited very poor antibody responses. A chimeric protein consisting of the receptor binding-deficient PA mutant tethered to the binding subunit of cholera toxin, which efficiently enters cells using the cholera toxin receptor rather than the PA receptor, elicited an anti-PA IgG antibody response similar to that elicited by wild-type PA; however, the chimeric protein elicited a poor toxin neutralizing antibody response. Taken together, our results demonstrate that the antigen capture pathway can dictate the magnitudes of the total IgG and toxin neutralizing antibody responses to PA as well as the ratio of the two responses.

Palytoxin: a new marine toxin from a coelenterate.

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Moore, R E; Scheuer, P J

1971-04-30

Palytoxin has been isolated from the zoanthids "limu-make-o-Hana" (Tentatively identified as Palythoa sp.) as a noncrystalline, chromatographically pure entity. Apart from polypeptide and protein toxins, it is the most highly toxic substance known, with a lethal dose (LD(59)) in mice of 0.15 microgram per kilogram by intravenous injection. Unlike the potent toxins batrachotoxin, saxitoxin, and tetrodotoxin which have molecular weights of 500 or less, palytoxin has an estimated molecular weight of 3300 and contains no repetitive amino acid or sugar units.

CD44 Promotes intoxication by the clostridial iota-family toxins.

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Wigelsworth, Darran J; Ruthel, Gordon; Schnell, Leonie; Herrlich, Peter; Blonder, Josip; Veenstra, Timothy D; Carman, Robert J; Wilkins, Tracy D; Van Nhieu, Guy Tran; Pauillac, Serge; Gibert, Maryse; Sauvonnet, Nathalie; Stiles, Bradley G; Popoff, Michel R; Barth, Holger

2012-01-01

Various pathogenic clostridia produce binary protein toxins associated with enteric diseases of humans and animals. Separate binding/translocation (B) components bind to a protein receptor on the cell surface, assemble with enzymatic (A) component(s), and mediate endocytosis of the toxin complex. Ultimately there is translocation of A component(s) from acidified endosomes into the cytosol, leading to destruction of the actin cytoskeleton. Our results revealed that CD44, a multifunctional surface protein of mammalian cells, facilitates intoxication by the iota family of clostridial binary toxins. Specific antibody against CD44 inhibited cytotoxicity of the prototypical Clostridium perfringens iota toxin. Versus CD44(+) melanoma cells, those lacking CD44 bound less toxin and were dose-dependently resistant to C. perfringens iota, as well as Clostridium difficile and Clostridium spiroforme iota-like, toxins. Purified CD44 specifically interacted in vitro with iota and iota-like, but not related Clostridium botulinum C2, toxins. Furthermore, CD44 knockout mice were resistant to iota toxin lethality. Collective data reveal an important role for CD44 during intoxication by a family of clostridial binary toxins.

Evaluation of Cholera Toxin Expression in Acidic, Alkaline and Neutral Conditions

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Narges Rahimi

2015-02-01

Full Text Available Background: Cholera is a severe disease which is caused by Vibrio cholerae and it is typically transmitted by either contaminated food or water particularly in developing countries. The most important virulence factor of this bacterium is an enterotoxin called cholera toxin which is a protein complex secreted by the Vibrio cholerae. Objectives: In this project, we determined the production of cholera toxin at different pH values. Materials and Methods: Two standard strain of Vibrio cholerae O1 biovar EL Tor N16961 and Vibrio cholerae O1 biovar Classic ATCC 14035 were used. After overnight cultivation of both the strains the total mRNA extracted and converted to total cDNA. Results: By Relative Real-Time PCR analysis the most cholera toxin production in classical and El Tor strains was at pH 8.5 and 8, respectively. Conclusions: Therefore, We may conclude that use of acidic diet will help in reduction of cholera toxin production.

Anthrax lethal toxin inhibits translation of hypoxia-inducible factor 1α and causes decreased tolerance to hypoxic stress.

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Ouyang, Weiming; Torigoe, Chikako; Fang, Hui; Xie, Tao; Frucht, David M

2014-02-14

Hypoxia is considered to be a contributor to the pathology associated with administration of anthrax lethal toxin (LT). However, we report here that serum lactate levels in LT-treated mice are reduced, a finding inconsistent with the anaerobic metabolism expected to occur during hypoxia. Reduced lactate levels are also observed in the culture supernatants of LT-treated cells. LT inhibits the accumulation of hypoxia-inducible factor (HIF)-1α, a subunit of HIF-1, the master regulator directing cellular responses to hypoxia. The toxin has no effect on the transcription or protein turnover of HIF-1α, but instead it acts to inhibit HIF-1α translation. LT treatment diminishes phosphorylation of eIF4B, eIF4E, and rpS6, critical components of the intracellular machinery required for HIF-1α translation. Moreover, blockade of MKK1/2-ERK1/2, but not p38 or JNK signaling, lowers HIF-1α protein levels in both normoxic and hypoxic conditions, consistent with a role for MKK1 and MKK2 as the major targets of LT responsible for the inhibition of HIF-1α translation. The physiological importance of the LT-induced translation blockade is demonstrated by the finding that LT treatment decreases the survival of hepatocyte cell lines grown in hypoxic conditions, an effect that is overcome by preinduction of HIF-1α. Taken together, these data support a role for LT in dysregulating HIF-1α and thereby disrupting homeostatic responses to hypoxia, an environmental characteristic of certain tissues at baseline and/or during disseminated infection with Bacillus anthracis.

A toxin-binding alkaline phosphatase fragment synergizes Bt toxin Cry1Ac against susceptible and resistant Helicoverpa armigera.

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Wenbo Chen

Full Text Available Evolution of resistance by insects threatens the continued success of pest control using insecticidal crystal (Cry proteins from the bacterium Bacillus thuringiensis (Bt in sprays and transgenic plants. In this study, laboratory selection with Cry1Ac yielded five strains of cotton bollworm, Helicoverpa armigera, with resistance ratios at the median lethal concentration (LC50 of activated Cry1Ac ranging from 22 to 1700. Reduced activity and reduced transcription of an alkaline phosphatase protein that binds Cry1Ac was associated with resistance to Cry1Ac in the four most resistant strains. A Cry1Ac-binding fragment of alkaline phosphatase from H. armigera (HaALP1f was not toxic by itself, but it increased mortality caused by Cry1Ac in a susceptible strain and in all five resistant strains. Although synergism of Bt toxins against susceptible insects by toxin-binding fragments of cadherin and aminopeptidase N has been reported previously, the results here provide the first evidence of synergism of a Bt toxin by a toxin-binding fragment of alkaline phosphatase. The results here also provide the first evidence of synergism of a Bt toxin by any toxin-binding peptide against resistant insects.

Binding properties of Clostridium botulinum type C progenitor toxin to mucins.

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Nakamura, Toshio; Takada, Noriko; Tonozuka, Takashi; Sakano, Yoshiyuki; Oguma, Keiji; Nishikawa, Atsushi

2007-04-01

It has been reported that Clostridium botulinum type C 16S progenitor toxin (C16S toxin) first binds to the sialic acid on the cell surface of mucin before invading cells [A. Nishikawa, N. Uotsu, H. Arimitsu, J.C. Lee, Y. Miura, Y. Fujinaga, H. Nakada, T. Watanabe, T. Ohyama, Y. Sakano, K. Oguma, The receptor and transporter for internalization of Clostridium botulinum type C progenitor toxin into HT-29 cells, Biochem. Biophys. Res. Commun. 319 (2004) 327-333]. In this study we investigated the binding properties of the C16S toxin to glycoproteins. Although the toxin bound to membrane blotted mucin derived from the bovine submaxillary gland (BSM), which contains a lot of sialyl oligosaccharides, it did not bind to neuraminidase-treated BSM. The binding of the toxin to BSM was inhibited by N-acetylneuraminic acid, N-glycolylneuraminic acid, and sialyl oligosaccharides strongly, but was not inhibited by neutral oligosaccharides. Both sialyl alpha2-3 lactose and sialyl alpha2-6 lactose prevented binding similarly. On the other hand, the toxin also bound well to porcine gastric mucin. In this case, neutral oligosaccharides might play an important role as ligand, since galactose and lactose inhibited binding. These results suggest that the toxin is capable of recognizing a wide variety of oligosaccharide structures.

Centrifugal microfluidic platform for ultrasensitive detection of Botulinum Toxin

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Botulinum neurotoxin – a global public health threat and category A bioterrorism agent - is the most toxic substance known and one of the most challenging toxins to detect due to its lethality at extremely low concentrations. Hence the live-mouse bioassay because of its superior sensitivity, remains...

Mass Spectrometric Identification and Differentiation of Botulinum Neurotoxins through Toxin Proteomics.

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Kalb, Suzanne R; Barr, John R

2013-08-01

Botulinum neurotoxins (BoNTs) cause the disease botulism, which can be lethal if untreated. There are seven known serotypes of BoNT, A-G, defined by their response to antisera. Many serotypes are distinguished into differing subtypes based on amino acid sequence and immunogenic properties, and some subtypes are further differentiated into toxin variants. Toxin characterization is important as different types of BoNT can respond differently to medical countermeasures for botulism, and characterization of the toxin can aid in epidemiologic and forensic investigations. Proteomic techniques have been established to determine the serotype, subtype, or toxin variant of BoNT. These techniques involve digestion of the toxin into peptides, tandem mass spectrometric (MS/MS) analysis of the peptides, and database searching to identify the BoNT protein. These techniques demonstrate the capability to detect BoNT and its neurotoxin-associated proteins, and differentiate the toxin from other toxins which are up to 99.9% identical in some cases. This differentiation can be accomplished from toxins present in a complex matrix such as stool, food, or bacterial cultures and no DNA is required.

Occurrence and sequestration of toxins in food chains.

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Mebs, D

1998-11-01

Animals may acquire toxicity by absorbing toxic compounds from their food, e.g. from plants or other animals. Sequestration and accumulation of toxins may provide protection from predators, which learn to avoid this prey because of unpleasant experiences such as bitter taste. This is a common phenomenon in marine as well as in terrestrial ecosystems. Moreover, toxins may enter food chains where they accumulate reaching high, often lethal concentrations. Palytoxin which had been primarily detected in marine zoanthids (Palythoa sp.), occurs also in a wide range of other animals, e.g. in sponges, corals, shellfish, polychaetes and crustaceans, but also in fish, which feed on crustaceans and zoanthids as well. These animals exhibit a high resistance to the toxin's action. The mechanisms which protect the Na+, K+-ATPase of their cell membranes, the primary target of palytoxin, is unknown. Sequestration of the toxin by other animals may cause health problems due to food poisoning.

Anthrax toxin: the long and winding road that leads to the kill.

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Abrami, Laurence; Reig, Nuria; van der Goot, F Gisou

2005-02-01

The past five years have led to a tremendous increase in our molecular understanding of the mode of action of the anthrax toxin, one of the two main virulence factors produced by Bacillus anthracis. The structures of each of the three components of the toxin--lethal factor (LF), edema factor (EF) and protective antigen (PA)--have been solved not only in their monomeric forms but, depending on the subunit, in a heptameric form, bound to their substrate, co-factor or receptor. The endocytic route followed by the toxin has also been unraveled and the enzymatic mechanisms of EF and LF elucidated.

Keeping the wolves at bay: antitoxins of prokaryotic type II toxin-antitoxin systems

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Wai Ting eChan

2016-03-01

Full Text Available In their initial stages of discovery, prokaryotic toxin-antitoxin (TA systems were confined to bacterial plasmids where they function to mediate the maintenance and stability of usually low- to medium-copy number plasmids through the post-segregational killing of any plasmid-free daughter cells that developed. Their eventual discovery as nearly ubiquitous and repetitive elements in bacterial chromosomes led to a wealth of knowledge and scientific debate as to their diversity and functionality in the prokaryotic lifestyle. Currently categorized into six different types designated types I – VI, type II TA systems are the best characterized. These generally comprised of two genes encoding a proteic toxin and its corresponding proteic antitoxin, respectively. Under normal growth conditions, the stable toxin is prevented from exerting its lethal effect through tight binding with the less stable antitoxin partner, forming a non-lethal TA protein complex. Besides binding with its cognate toxin, the antitoxin also plays a role in regulating the expression of the type II TA operon by binding to the operator site, thereby repressing transcription from the TA promoter. In most cases, full repression is observed in the presence of the TA complex as binding of the toxin enhances the DNA binding capability of the antitoxin. TA systems have been implicated in a gamut of prokaryotic cellular functions such as being mediators of programmed cell death as well as persistence or dormancy, biofilm formation, as defensive weapons against bacteriophage infections and as virulence factors in pathogenic bacteria. It is thus apparent that these antitoxins, as DNA-binding proteins, play an essential role in modulating the prokaryotic lifestyle whilst at the same time preventing the lethal action of the toxins under normal growth conditions, i.e., keeping the proverbial wolves at bay. In this review, we will cover the diversity and characteristics of various type II TA

Inhibition of Clostridium difficile toxin A and B by 1,2-cyclohexanedione modification of an arginine residue.

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Balfanz, J; Rautenberg, P

1989-12-29

Toxin A (enterotoxin) and toxin B (cytotoxin) of Clostridium difficile were both inactivated by the arginine specific reagent 1,2-cyclohexanedione. Molecular stability during the inactivation process was demonstrated by SDS-PAGE analysis showing the same migration rates for modified and unmodified forms of the 230 kDa toxin A and of the 250 kDa toxin B. Cytotoxicity of both toxins as well as mouse lethality of the enterotoxin were drastically decreased as a result of the arginine modification. The reaction followed pseudo-first-order kinetics. Analysis of the data suggested that modification of a single arginine residue was sufficient to abolish the activity of both toxins.

Structure-function analyses reveal the molecular architecture and neutralization mechanism of a bacterial HEPN-MNT toxin-antitoxin system.

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Jia, Xuanyan; Yao, Jianyun; Gao, Zengqiang; Liu, Guangfeng; Dong, Yu-Hui; Wang, Xiaoxue; Zhang, Heng

2018-05-04

Toxin-antitoxin (TA) loci in bacteria are small genetic modules that regulate various cellular activities, including cell growth and death. The two-gene module encoding a HEPN (higher eukaryotes and prokaryotes nucleotide-binding) domain and a cognate MNT (minimal nucleotidyltransferase) domain have been predicted to represent a novel type II TA system prevalent in archaea and bacteria. However, the neutralization mechanism and cellular targets of the TA family remain unclear. The toxin SO_3166 having a HEPN domain and its cognate antitoxin SO_3165 with an MNT domain constitute a typical type II TA system that regulates cell motility and confers plasmid stability in the bacterium Shewanella oneidensis Here, we report the crystal structure and solution conformation of the SO_3166-SO_3165 pair, representing the first complex structures in this TA family. The structures revealed that SO_3165 and SO_3166 form a tight heterooctamer (at a 2:6 ratio), an organization that is very rare in other TA systems. We also observed that SO_3166 dimerization enables the formation of a deep cleft at the HEPN-domain interface harboring a composite R X 4-6H active site that functions as an RNA-cleaving RNase. SO_3165 bound SO_3166 mainly through its two α-helices (α2 and α4), functioning as molecular recognition elements. Moreover, their insertion into the SO_3166 cleft sterically blocked the R X 4-6H site or narrowed the cleft to inhibit RNA substrate binding. Structure-based mutagenesis confirmed the important roles of these α-helices in SO_3166 binding and inhibition. Our structure-function analysis provides first insights into the neutralization mechanism of the HEPN-MNT TA family. © 2018 Jia et al.

Staphylococcus aureus α-Toxin: Nearly a Century of Intrigue

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Bryan J. Berube

2013-06-01

Full Text Available Staphylococcus aureus secretes a number of host-injurious toxins, among the most prominent of which is the small β-barrel pore-forming toxin α-hemolysin. Initially named based on its properties as a red blood cell lytic toxin, early studies suggested a far greater complexity of α-hemolysin action as nucleated cells also exhibited distinct responses to intoxication. The hemolysin, most aptly referred to as α-toxin based on its broad range of cellular specificity, has long been recognized as an important cause of injury in the context of both skin necrosis and lethal infection. The recent identification of ADAM10 as a cellular receptor for α-toxin has provided keen insight on the biology of toxin action during disease pathogenesis, demonstrating the molecular mechanisms by which the toxin causes tissue barrier disruption at host interfaces lined by epithelial or endothelial cells. This review highlights both the historical studies that laid the groundwork for nearly a century of research on α-toxin and key findings on the structural and functional biology of the toxin, in addition to discussing emerging observations that have significantly expanded our understanding of this toxin in S. aureus disease. The identification of ADAM10 as a proteinaceous receptor for the toxin not only provides a greater appreciation of truths uncovered by many historic studies, but now affords the opportunity to more extensively probe and understand the role of α-toxin in modulation of the complex interaction of S. aureus with its human host.

Selective effects of an octopus toxin on action potentials

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Dulhunty, Angela; Gage, Peter W.

1971-01-01

1. A lethal, water soluble toxin (Maculotoxin, MTX) with a molecular weight less than 540, can be extracted from the salivary glands of an octopus (Hapalochlaena maculosa). 2. MTX blocks action potentials in sartorius muscle fibres of toads without affecting the membrane potential. Delayed rectification is not inhibited by the toxin. 3. At low concentrations (10-6-10-5 g/ml.) MTX blocks action potentials only after a certain number have been elicited. The number of action potentials, which can be defined accurately, depends on the concentration of MTX and the concentration of sodium ions in the extracellular solution. 4. The toxin has no post-synaptic effect at the neuromuscular junction and it is concluded that it blocks neuromuscular transmission by inhibiting action potentials in motor nerve terminals. PMID:4330930

Tityus serrulatus venom--A lethal cocktail.

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Pucca, Manuela Berto; Cerni, Felipe Augusto; Pinheiro Junior, Ernesto Lopes; Bordon, Karla de Castro Figueiredo; Amorim, Fernanda Gobbi; Cordeiro, Francielle Almeida; Longhim, Heloisa Tavoni; Cremonez, Caroline Marroni; Oliveira, Guilherme Honda; Arantes, Eliane Candiani

2015-12-15

Tityus serrulatus (Ts) is the main scorpion species of medical importance in Brazil. Ts venom is composed of several compounds such as mucus, inorganic salts, lipids, amines, nucleotides, enzymes, kallikrein inhibitor, natriuretic peptide, proteins with high molecular mass, peptides, free amino acids and neurotoxins. Neurotoxins are considered the most responsible for the envenoming syndrome due to their pharmacological action on ion channels such as voltage-gated sodium (Nav) and potassium (Kv) channels. The major goal of this review is to present important advances in Ts envenoming research, correlating both the crude Ts venom and isolated toxins with alterations observed in all human systems. The most remarkable event lies in the Ts induced massive releasing of neurotransmitters influencing, directly or indirectly, the entire body. Ts venom proved to extremely affect nervous and muscular systems, to modulate the immune system, to induce cardiac disorders, to cause pulmonary edema, to decrease urinary flow and to alter endocrine, exocrine, reproductive, integumentary, skeletal and digestive functions. Therefore, Ts venom possesses toxins affecting all anatomic systems, making it a lethal cocktail. However, its low lethality may be due to the low venom mass injected, to the different venom compositions, the body characteristics and health conditions of the victim and the local of Ts sting. Furthermore, we also described the different treatments employed during envenoming cases. In particular, throughout the review, an effort will be made to provide information from an extensive documented studies concerning Ts venom in vitro, in animals and in humans (a total of 151 references). Copyright © 2015 Elsevier Ltd. All rights reserved.

Lipoproteins/peptides are sepsis-inducing toxins from bacteria that can be neutralized by synthetic anti-endotoxin peptides.

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Martinez de Tejada, Guillermo; Heinbockel, Lena; Ferrer-Espada, Raquel; Heine, Holger; Alexander, Christian; Bárcena-Varela, Sergio; Goldmann, Torsten; Correa, Wilmar; Wiesmüller, Karl-Heinz; Gisch, Nicolas; Sánchez-Gómez, Susana; Fukuoka, Satoshi; Schürholz, Tobias; Gutsmann, Thomas; Brandenburg, Klaus

2015-09-22

Sepsis, a life-threatening syndrome with increasing incidence worldwide, is triggered by an overwhelming inflammation induced by microbial toxins released into the bloodstream during infection. A well-known sepsis-inducing factor is the membrane constituent of Gram-negative bacteria, lipopolysaccharide (LPS), signalling via Toll-like receptor-4. Although sepsis is caused in more than 50% cases by Gram-positive and mycoplasma cells, the causative compounds are still poorly described. In contradicting investigations lipoproteins/-peptides (LP), lipoteichoic acids (LTA), and peptidoglycans (PGN), were made responsible for eliciting this pathology. Here, we used human mononuclear cells from healthy donors to determine the cytokine-inducing activity of various LPs from different bacterial origin, synthetic and natural, and compared their activity with that of natural LTA and PGN. We demonstrate that LP are the most potent non-LPS pro-inflammatory toxins of the bacterial cell walls, signalling via Toll-like receptor-2, not only in vitro, but also when inoculated into mice: A synthetic LP caused sepsis-related pathological symptoms in a dose-response manner. Additionally, these mice produced pro-inflammatory cytokines characteristic of a septic reaction. Importantly, the recently designed polypeptide Aspidasept(®) which has been proven to efficiently neutralize LPS in vivo, inhibited cytokines induced by the various non-LPS compounds protecting animals from the pro-inflammatory activity of synthetic LP.

A Supercluster of Neutralizing Epitopes at the Interface of Ricin’s Enzymatic (RTA and Binding (RTB Subunits

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Amanda Y. Poon

2017-11-01

Full Text Available As part of an effort to engineer ricin antitoxins and immunotherapies, we previously produced and characterized a collection of phage-displayed, heavy chain-only antibodies (VHHs from alpacas that had been immunized with ricin antigens. In our initial screens, we identified nine VHHs directed against ricin toxin’s binding subunit (RTB, but only one, JIZ-B7, had toxin-neutralizing activity. Linking JIZ-B7 to different VHHs against ricin’s enzymatic subunit (RTA resulted in several bispecific antibodies with potent toxin-neutralizing activity in vitro and in vivo. JIZ-B7 may therefore be an integral component of a future VHH-based neutralizing agent (VNA for ricin toxin. In this study, we now localize, using competitive ELISA, JIZ-B7’s epitope to a region of RTB’s domain 2 sandwiched between the high-affinity galactose/N-acetylgalactosamine (Gal/GalNAc-binding site and the boundary of a neutralizing hotspot on RTA known as cluster II. Analysis of additional RTB (n = 8- and holotoxin (n = 4-specific VHHs from a recent series of screens identified a “supercluster” of neutralizing epitopes at the RTA-RTB interface. Among the VHHs tested, toxin-neutralizing activity was most closely associated with epitope proximity to RTA, and not interference with RTB’s ability to engage Gal/GalNAc receptors. We conclude that JIZ-B7 is representative of a larger group of potent toxin-neutralizing antibodies, possibly including many described in the literature dating back several decades, that recognize tertiary and possibly quaternary epitopes located at the RTA-RTB interface and that target a region of vulnerability on ricin toxin.

Proteinaceous toxins from three species of scorpaeniform fish (lionfish Pterois lunulata, devil stinger Inimicus japonicus and waspfish Hypodytes rubripinnis): close similarity in properties and primary structures to stonefish toxins.

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Kiriake, Aya; Suzuki, Yasuko; Nagashima, Yuji; Shiomi, Kazuo

2013-08-01

The crude toxins from three species of venomous fish (lionfish Pterois lunulata, devil stinger Inimicus japonicus and waspfish Hypodytes rubripinnis) belonging to the order Scorpaeniformes exhibited mouse-lethal, hemolytic, edema-forming and nociceptive activities. In view of the antigenic cross-reactivity with the stonefish toxins, the primary structures of the stonefish toxin-like toxins from the three scorpaeniform fish were determined by cDNA cloning using primers designed from the highly conserved sequences of the stonefish toxins. Based on the data obtained in gel filtration, immunoblotting and cDNA cloning, each toxin was judged to be a 160 kDa heterodimer composed of 80 kDa α- and β-subunits. The three scorpaeniform fish toxins contain a B30.2/SPRY domain (∼200 amino acid residues) in the C-terminal region of each subunit, as reported for the toxins from two species of lionfish and two species of stonefish. With respect to the amino acid sequence similarity, the scorpaeniform fish toxins are divided into the following two groups: toxins from three species of lionfish and those from devil stinger, two species of stonefish and waspfish. The phylogenetic tree generated also clearly supports the classification of the toxins. Copyright © 2013 Elsevier Ltd. All rights reserved.

An insecticidal toxin from Nephila clavata spider venom.

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Jin, Lin; Fang, Mingqian; Chen, Mengrou; Zhou, Chunling; Ombati, Rose; Hakim, Md Abdul; Mo, Guoxiang; Lai, Ren; Yan, Xiuwen; Wang, Yumin; Yang, Shilong

2017-07-01

Spiders are the most successful insect predators given that they use their venom containing insecticidal peptides as biochemical weapons for preying. Due to the high specificity and potency of peptidic toxins, discoveries of insecticidal toxins from spider venom have provided an opportunity to obtain natural compounds for agricultural applications without affecting human health. In this study, a novel insecticidal toxin (μ-NPTX-Nc1a) was identified and characterized from the venom of Nephila clavata. Its primary sequence is GCNPDCTGIQCGWPRCPGGQNPVMDKCVSCCPFCPPKSAQG which was determined by automated Edman degradation, cDNA cloning, and MS/MS analysis. BLAST search indicated that Nc1a shows no similarity with known peptides or proteins, indicating that Nc1a belongs to a novel family of insecticidal peptide. Nc1a displayed inhibitory effects on Na V and K V channels in cockroach dorsal unpaired median neurons. The median lethal dose (LD50) of Nc1a on cockroach was 573 ng/g. Herein, a study that identifies a novel insecticidal toxin, which can be a potential candidate and/or template for the development of bioinsecticides, is presented.

Identification and characterization of B-cell epitopes of 3FTx and PLA(2) toxins from Micrurus corallinus snake venom.

Science.gov (United States)

Castro, K L; Duarte, C G; Ramos, H R; Machado de Avila, R A; Schneider, F S; Oliveira, D; Freitas, C F; Kalapothakis, E; Ho, P L; Chávez-Olortegui, C

2015-01-01

The main goal of this work was to develop a strategy to identify B-cell epitopes on four different three finger toxins (3FTX) and one phospholipase A2 (PLA2) from Micrurus corallinus snake venom. 3FTx and PLA2 are highly abundant components in Elapidic venoms and are the major responsibles for the toxicity observed in envenomation by coral snakes. Overlapping peptides from the sequence of each toxin were prepared by SPOT method and three different anti-elapidic sera were used to map the epitopes. After immunogenicity analysis of the spot-reactive peptides by EPITOPIA, a computational method, nine sequences from the five toxins were chemically synthesized and antigenically and immunogenically characterized. All the peptides were used together as immunogens in rabbits, delivered with Freund's adjuvant for a first cycle of immunization and Montanide in the second. A good antibody response against individual synthetic peptides and M. corallinus venom was achieved. Anti-peptide IgGs were also cross-reactive against Micrurus frontalis and Micrurus lemniscatus crude venoms. In addition, anti-peptide IgGs inhibits the lethal and phospholipasic activities of M. corallinus crude venom. Our results provide a rational basis to the identification of neutralizing epitopes on coral snake toxins and show that their corresponding synthetic peptides could improve the generation of immuno-therapeutics. The use of synthetic peptide for immunization is a reasonable approach, since it enables poly-specificity, low risk of toxic effects and large scale production. Copyright © 2014 Elsevier Ltd. All rights reserved.

Dengue virus specific IgY provides protection following lethal dengue virus challenge and is neutralizing in the absence of inducing antibody dependent enhancement.

Science.gov (United States)

Fink, Ashley L; Williams, Katherine L; Harris, Eva; Alvine, Travis D; Henderson, Thomas; Schiltz, James; Nilles, Matthew L; Bradley, David S

2017-07-01

Dengue hemorrhagic fever (DHF) and dengue shock syndrome (DSS) are severe disease manifestations that can occur following sequential infection with different dengue virus serotypes (DENV1-4). At present, there are no licensed therapies to treat DENV-induced disease. DHF and DSS are thought to be mediated by serotype cross-reactive antibodies that facilitate antibody-dependent enhancement (ADE) by binding to viral antigens and then Fcγ receptors (FcγR) on target myeloid cells. Using genetically engineered DENV-specific antibodies, it has been shown that the interaction between the Fc portion of serotype cross-reactive antibodies and FcγR is required to induce ADE. Additionally, it was demonstrated that these antibodies were as neutralizing as their non-modified variants, were incapable of inducing ADE, and were therapeutic following a lethal, antibody-enhanced infection. Therefore, we hypothesized that avian IgY, which do not interact with mammalian FcγR, would provide a novel therapy for DENV-induced disease. We demonstrate here that goose-derived anti-DENV2 IgY neutralized DENV2 and did not induce ADE in vitro. Anti-DENV2 IgY was also protective in vivo when administered 24 hours following a lethal DENV2 infection. We were also able to demonstrate via epitope mapping that both full-length and alternatively spliced anti-DENV2 IgY recognized different epitopes, including epitopes that have not been previously identified. These observations provide evidence for the potential therapeutic applications of goose-derived anti-DENV2 IgY.

Dengue virus specific IgY provides protection following lethal dengue virus challenge and is neutralizing in the absence of inducing antibody dependent enhancement.

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Ashley L Fink

2017-07-01

Full Text Available Dengue hemorrhagic fever (DHF and dengue shock syndrome (DSS are severe disease manifestations that can occur following sequential infection with different dengue virus serotypes (DENV1-4. At present, there are no licensed therapies to treat DENV-induced disease. DHF and DSS are thought to be mediated by serotype cross-reactive antibodies that facilitate antibody-dependent enhancement (ADE by binding to viral antigens and then Fcγ receptors (FcγR on target myeloid cells. Using genetically engineered DENV-specific antibodies, it has been shown that the interaction between the Fc portion of serotype cross-reactive antibodies and FcγR is required to induce ADE. Additionally, it was demonstrated that these antibodies were as neutralizing as their non-modified variants, were incapable of inducing ADE, and were therapeutic following a lethal, antibody-enhanced infection. Therefore, we hypothesized that avian IgY, which do not interact with mammalian FcγR, would provide a novel therapy for DENV-induced disease. We demonstrate here that goose-derived anti-DENV2 IgY neutralized DENV2 and did not induce ADE in vitro. Anti-DENV2 IgY was also protective in vivo when administered 24 hours following a lethal DENV2 infection. We were also able to demonstrate via epitope mapping that both full-length and alternatively spliced anti-DENV2 IgY recognized different epitopes, including epitopes that have not been previously identified. These observations provide evidence for the potential therapeutic applications of goose-derived anti-DENV2 IgY.

Robustness against serum neutralization of a poliovirus type 1 from a lethal epidemic of poliomyelitis in the Republic of Congo in 2010.

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Drexler, Jan Felix; Grard, Gilda; Lukashev, Alexander N; Kozlovskaya, Liubov I; Böttcher, Sindy; Uslu, Gökhan; Reimerink, Johan; Gmyl, Anatoly P; Taty-Taty, Raphaël; Lekana-Douki, Sonia Etenna; Nkoghe, Dieudonné; Eis-Hübinger, Anna M; Diedrich, Sabine; Koopmans, Marion; Leroy, Eric M; Drosten, Christian

2014-09-02

In 2010, a large outbreak of poliomyelitis with unusual 47% lethality occurred in Pointe Noire, Republic of Congo. Vaccine-mediated immunity against the outbreak virus was never investigated. A wild poliovirus 1 (WPV1) isolated from a fatal case (termed PV1-RC2010) showed a previously unknown combination of amino acid exchanges in critical antigenic site 2 (AgS2, VP1 capsid protein positions 221SAAL → 221PADL). These exchanges were also detected in an additional 11 WPV1 strains from fatal cases. PV1-RC2010 escaped neutralization by three different mAbs relevant for AgS2. Virus neutralization was tested in sera from fatal cases, who died before supplementary immunization (n = 24), Gabonese recipients of recent oral polio vaccination (n = 12), routinely vaccinated German medical students (n = 34), and German outpatients tested for antipoliovirus immunity (n = 17) on Vero, human rhabdomyosarcoma, and human epidermoid carcinoma 2 cells. Fatal poliomyelitis cases gave laboratory evidence of previous trivalent vaccination. Neutralizing antibody titers against PV1-RC2010 were significantly lower than those against the vaccine strain Sabin-1, two genetically distinct WPV1s isolated in 1965 and 2010 and two genetically distinct vaccine-derived PV strains. Of German vaccinees tested according to World Health Organization protocols, 15-29% were unprotected according to their neutralization titers (poliomyelitis eradication in populations with predominantly vaccine-derived immunity. Sustained vaccination coverage and clinical and environmental surveillance will be necessary.

Specific egg yolk immunoglobulin as a new preventive approach for Shiga-toxin-mediated diseases.

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Paola Neri

Full Text Available Shiga toxins (Stxs are involved in the development of severe systemic complications associated with enterohemorrhagic Escherichia coli (EHEC infection. Various neutralizing agents against Stxs are under investigation for management of EHEC infection. In this study, we immunized chickens with formalin-inactivated Stx-1 or Stx-2, and obtained immunoglobulin Y (IgY from the egg yolk. Anti-Stx-1 IgY and anti-Stx-2 IgY recognized the corresponding Stx A subunit and polymeric but not monomeric B subunit. Anti-Stx-1 IgY and anti-Stx-2 IgY suppressed the cytotoxicity of Stx-1 and Stx-2 to HeLa 229 cells, without cross-suppressive activity. The suppressive activity of these IgY was abrogated by pre-incubation with the corresponding recombinant B subunit, which suggests that the antibodies directed to the polymeric B subunits were predominantly involved in the suppression. In vivo, the intraperitoneal or intravenous administration of these IgY rescued mice from death caused by intraperitoneal injection of the corresponding toxin at a lethal dose. Moreover, oral administration of anti-Stx-2 IgY reduced the mortality of mice infected intestinally with EHEC O157:H7. Our results therefore suggest that anti-Stx IgY antibodies may be considered as preventive agents for Stx-mediated diseases in EHEC infection.

Semicarbazone EGA Inhibits Uptake of Diphtheria Toxin into Human Cells and Protects Cells from Intoxication

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Leonie Schnell

2016-07-01

Full Text Available Diphtheria toxin is a single-chain protein toxin that invades human cells by receptor-mediated endocytosis. In acidic endosomes, its translocation domain inserts into endosomal membranes and facilitates the transport of the catalytic domain (DTA from endosomal lumen into the host cell cytosol. Here, DTA ADP-ribosylates elongation factor 2 inhibits protein synthesis and leads to cell death. The compound 4-bromobenzaldehyde N-(2,6-dimethylphenylsemicarbazone (EGA has been previously shown to protect cells from various bacterial protein toxins which deliver their enzymatic subunits from acidic endosomes to the cytosol, including Bacillus anthracis lethal toxin and the binary clostridial actin ADP-ribosylating toxins C2, iota and Clostridium difficile binary toxin (CDT. Here, we demonstrate that EGA also protects human cells from diphtheria toxin by inhibiting the pH-dependent translocation of DTA across cell membranes. The results suggest that EGA might serve for treatment and/or prevention of the severe disease diphtheria.

Therapeutic efficacy of antibodies lacking Fcγ receptor binding against lethal dengue virus infection is due to neutralizing potency and blocking of enhancing antibodies [corrected].

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Katherine L Williams

2013-02-01

Full Text Available Dengue hemorrhagic fever and dengue shock syndrome (DHF/DSS are life-threatening complications following infection with one of the four serotypes of dengue virus (DENV. At present, no vaccine or antiviral therapies are available against dengue. Here, we characterized a panel of eight human or mouse-human chimeric monoclonal antibodies (MAbs and their modified variants lacking effector function and dissected the mechanism by which some protect against antibody-enhanced lethal DENV infection. We found that neutralizing modified MAbs that recognize the fusion loop or the A strand epitopes on domains II and III of the envelope protein, respectively, act therapeutically by competing with and/or displacing enhancing antibodies. By analyzing these relationships, we developed a novel in vitro suppression-of-enhancement assay that predicts the ability of modified MAbs to act therapeutically against antibody-enhanced disease in vivo. These studies provide new insight into the biology of DENV pathogenesis and the requirements for antibodies to treat lethal DENV disease.

Staphylococcus aureus β-Toxin Mutants Are Defective in Biofilm Ligase and Sphingomyelinase Activity, and Causation of Infective Endocarditis and Sepsis.

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Herrera, Alfa; Vu, Bao G; Stach, Christopher S; Merriman, Joseph A; Horswill, Alexander R; Salgado-Pabón, Wilmara; Schlievert, Patrick M

2016-05-03

β-Toxin is an important virulence factor of Staphylococcus aureus, contributing to colonization and development of disease [Salgado-Pabon, W., et al. (2014) J. Infect. Dis. 210, 784-792; Huseby, M. J., et al. (2010) Proc. Natl. Acad. Sci. U.S.A. 107, 14407-14412; Katayama, Y., et al. (2013) J. Bacteriol. 195, 1194-1203]. This cytotoxin has two distinct mechanisms of action: sphingomyelinase activity and DNA biofilm ligase activity. However, the distinct mechanism that is most important for its role in infective endocarditis is unknown. We characterized the active site of β-toxin DNA biofilm ligase activity by examining deficiencies in site-directed mutants through in vitro DNA precipitation and biofilm formation assays. Possible conformational changes in mutant structure compared to that of wild-type toxin were assessed preliminarily by trypsin digestion analysis, retention of sphingomyelinase activity, and predicted structures based on the native toxin structure. We addressed the contribution of each mechanism of action to producing infective endocarditis and sepsis in vivo in a rabbit model. The H289N β-toxin mutant, lacking sphingomyelinase activity, exhibited lower sepsis lethality and infective endocarditis vegetation formation compared to those of the wild-type toxin. β-Toxin mutants with disrupted biofilm ligase activity did not exhibit decreased sepsis lethality but were deficient in infective endocarditis vegetation formation compared to the wild-type protein. Our study begins to characterize the DNA biofilm ligase active site of β-toxin and suggests β-toxin functions importantly in infective endocarditis through both of its mechanisms of action.

Antivenom Cross-Neutralization of the Venoms of Hydrophis schistosus and Hydrophis curtus, Two Common Sea Snakes in Malaysian Waters

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Choo Hock Tan

2015-02-01

Full Text Available Sea snake envenomation is a serious occupational hazard in tropical waters. In Malaysia, the beaked sea snake (Hydrophis schistosus, formerly known as Enhydrina schistosa and the spine-bellied sea snake (Hydrophis curtus, formerly known as Lapemis curtus or Lapemis hardwickii are two commonly encountered species. Australian CSL sea snake antivenom is the definitive treatment for sea snake envenomation; it is unfortunately extremely costly locally and is not widely available or adequately stocked in local hospitals. This study investigated the cross-neutralizing potential of three regionally produced anti-cobra antivenoms against the venoms of Malaysian H. schistosus and H. curtus. All three antivenoms conferred paraspecific protection from sea snake venom lethality in mice, with potency increasing in the following order: Taiwan bivalent antivenom < Thai monocled cobra monovalent antivenom < Thai neuro polyvalent antivenom (NPAV. NPAV demonstrated cross-neutralizing potencies of 0.4 mg/vial for H. schistosus venom and 0.8 mg/vial for H. curtus, which translates to a dose of less than 20 vials of NPAV to neutralize an average amount of sea snake venom per bite (inferred from venom milking. The cross-neutralization activity was supported by ELISA cross-reactivity between NPAV and the venoms of H. schistosus (58.4% and H. curtus (70.4%. These findings revealed the potential of NPAV as a second-line treatment for sea snake envenomation in the region. Further profiling of the cross-neutralization activity should address the antivenomic basis using purified toxin-based assays.

A Viral Nanoparticle with Dual Function as an Anthrax Antitoxin and Vaccine

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Manayani, Darly J; Thomas, Diane; Dryden, Kelly A; Reddy, Vijay; Siladi, Marc E; Marlett, John M; Rainey, G. Jonah A; Pique, Michael E; Scobie, Heather M; Yeager, Mark; Young, John A. T; Manchester, Marianne; Schneemann, Anette

2007-01-01

The recent use of Bacillus anthracis as a bioweapon has stimulated the search for novel antitoxins and vaccines that act rapidly and with minimal adverse effects. B. anthracis produces an AB-type toxin composed of the receptor-binding moiety protective antigen (PA) and the enzymatic moieties edema factor and lethal factor. PA is a key target for both antitoxin and vaccine development. We used the icosahedral insect virus Flock House virus as a platform to display 180 copies of the high affinity, PA-binding von Willebrand A domain of the ANTXR2 cellular receptor. The chimeric virus-like particles (VLPs) correctly displayed the receptor von Willebrand A domain on their surface and inhibited lethal toxin action in in vitro and in vivo models of anthrax intoxication. Moreover, VLPs complexed with PA elicited a potent toxin-neutralizing antibody response that protected rats from anthrax lethal toxin challenge after a single immunization without adjuvant. This recombinant VLP platform represents a novel and highly effective, dually-acting reagent for treatment and protection against anthrax. PMID:17922572

Brown spider dermonecrotic toxin directly induces nephrotoxicity

International Nuclear Information System (INIS)

Chaim, Olga Meiri; Sade, Youssef Bacila; Bertoni da Silveira, Rafael; Toma, Leny; Kalapothakis, Evanguedes; Chavez-Olortegui, Carlos; Mangili, Oldemir Carlos; Gremski, Waldemiro; Dietrich, Carl Peter von; Nader, Helena B.; Sanches Veiga, Silvio

2006-01-01

culture substratum. In addition, dermonecrotic toxin treatment of MDCK cells changed their viability evaluated by XTT and Neutral-Red Uptake methodologies. The present results point to brown spider dermonecrotic toxin cytotoxicity upon renal structures in vivo and renal cells in vitro and provide experimental evidence that this brown spider toxin is directly involved in nephrotoxicity evoked during Loxosceles spider venom accidents

The lethality test used for estimating the potency of antivenoms against Bothrops asper snake venom: pathophysiological mechanisms, prophylactic analgesia, and a surrogate in vitro assay.

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Chacón, Francisco; Oviedo, Andrea; Escalante, Teresa; Solano, Gabriela; Rucavado, Alexandra; Gutiérrez, José María

2015-01-01

The potency of antivenoms is assessed by analyzing the neutralization of venom-induced lethality, and is expressed as the Median Effective Dose (ED50). The present study was designed to investigate the pathophysiological mechanisms responsible for lethality induced by the venom of Bothrops asper, in the experimental conditions used for the evaluation of the neutralizing potency of antivenoms. Mice injected with 4 LD50s of venom by the intraperitoneal route died within ∼25 min with drastic alterations in the abdominal organs, characterized by hemorrhage, increment in plasma extravasation, and hemoconcentration, thus leading to hypovolemia and cardiovascular collapse. Snake venom metalloproteinases (SVMPs) play a predominat role in lethality, as judged by partial inhibition by the chelating agent CaNa2EDTA. When venom was mixed with antivenom, there was a venom/antivenom ratio at which hemorrhage was significantly reduced, but mice died at later time intervals with evident hemoconcentration, indicating that other components in addition to SVMPs also contribute to plasma extravasation and lethality. Pretreatment with the analgesic tramadol did not affect the outcome of the neutralization test, thus suggesting that prophylactic (precautionary) analgesia can be introduced in this assay. Neutralization of lethality in mice correlated with neutralization of in vitro coagulant activity in human plasma. Copyright © 2014 Elsevier Ltd. All rights reserved.

A Medical Research and Evaluation Facility (MREF) and Studies Supporting the Medical Chemical Defense Program: Task 95-39: Methods Development and Validation of Two Mouse Bioassays for Use in Quantifying Botulinum Toxins (A, B, C, D and E) and Toxin Antibody Titers

National Research Council Canada - National Science Library

Olson, Carl

1997-01-01

Ths task was conducted for the U.S. Army Medical Materiel Development Activity (USAMMDA) to validate two mouse bioassays for quantify botulinum toxin potency and neutralizing antibodies to botulimun toxins...

Recombinant Protein Containing B-Cell Epitopes of Different Loxosceles Spider Toxins Generates Neutralizing Antibodies in Immunized Rabbits.

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Lima, Sabrina de Almeida; Guerra-Duarte, Clara; Costal-Oliveira, Fernanda; Mendes, Thais Melo; Figueiredo, Luís F M; Oliveira, Daysiane; Machado de Avila, Ricardo A; Ferrer, Valéria Pereira; Trevisan-Silva, Dilza; Veiga, Silvio S; Minozzo, João C; Kalapothakis, Evanguedes; Chávez-Olórtegui, Carlos

2018-01-01

Loxoscelism is the most important form of araneism in South America. The treatment of these accidents uses heterologous antivenoms obtained from immunization of production animals with crude loxoscelic venom. Due to the scarcity of this immunogen, new alternatives for its substitution in antivenom production are of medical interest. In the present work, three linear epitopes for Loxosceles astacin-like protease 1 (LALP-1) (SLGRGCTDFGTILHE, ENNTRTIGPFDYDSIMLYGAY, and KLYKCPPVNPYPGGIRPYVNV) and two for hyaluronidase (LiHYAL) (NGGIPQLGDLKAHLEKSAVDI and ILDKSATGLRIIDWEAWR) from Loxosceles intermedia spider venom were identified by SPOT-synthesis technique. One formerly characterized linear epitope (DFSGPYLPSLPTLDA) of sphingomyelinase D (SMase D) SMase-I from Loxosceles laeta was also chosen to constitute a new recombinant multiepitopic protein. These epitopes were combined with a previously produced chimeric multiepitopic protein (rCpLi) composed by linear and conformational B-cell epitopes from SMase D from L. intermedia venom, generating a new recombinant multiepitopic protein derived from loxoscelic toxins (rMEPLox). We demonstrated that rMEPLox is non-toxic and antibodies elicited in rabbits against this antigen present reactivity in ELISA and immunoblot assays with Brazilian L. intermedia, L. laeta, L. gaucho , and L. similis spider venoms. In vivo and in vitro neutralization assays showed that anti-rMEPLox antibodies can efficiently neutralize the sphingomyelinase, hyaluronidase, and metalloproteinase activity of L. intermedia venom. This study suggests that this multiepitopic protein can be a suitable candidate for experimental vaccination approaches or for antivenom production against Loxosceles spp. venoms.

Recombinant Protein Containing B-Cell Epitopes of Different Loxosceles Spider Toxins Generates Neutralizing Antibodies in Immunized Rabbits

Science.gov (United States)

Lima, Sabrina de Almeida; Guerra-Duarte, Clara; Costal-Oliveira, Fernanda; Mendes, Thais Melo; Figueiredo, Luís F. M.; Oliveira, Daysiane; Machado de Avila, Ricardo A.; Ferrer, Valéria Pereira; Trevisan-Silva, Dilza; Veiga, Silvio S.; Minozzo, João C.; Kalapothakis, Evanguedes; Chávez-Olórtegui, Carlos

2018-01-01

Loxoscelism is the most important form of araneism in South America. The treatment of these accidents uses heterologous antivenoms obtained from immunization of production animals with crude loxoscelic venom. Due to the scarcity of this immunogen, new alternatives for its substitution in antivenom production are of medical interest. In the present work, three linear epitopes for Loxosceles astacin-like protease 1 (LALP-1) (SLGRGCTDFGTILHE, ENNTRTIGPFDYDSIMLYGAY, and KLYKCPPVNPYPGGIRPYVNV) and two for hyaluronidase (LiHYAL) (NGGIPQLGDLKAHLEKSAVDI and ILDKSATGLRIIDWEAWR) from Loxosceles intermedia spider venom were identified by SPOT-synthesis technique. One formerly characterized linear epitope (DFSGPYLPSLPTLDA) of sphingomyelinase D (SMase D) SMase-I from Loxosceles laeta was also chosen to constitute a new recombinant multiepitopic protein. These epitopes were combined with a previously produced chimeric multiepitopic protein (rCpLi) composed by linear and conformational B-cell epitopes from SMase D from L. intermedia venom, generating a new recombinant multiepitopic protein derived from loxoscelic toxins (rMEPLox). We demonstrated that rMEPLox is non-toxic and antibodies elicited in rabbits against this antigen present reactivity in ELISA and immunoblot assays with Brazilian L. intermedia, L. laeta, L. gaucho, and L. similis spider venoms. In vivo and in vitro neutralization assays showed that anti-rMEPLox antibodies can efficiently neutralize the sphingomyelinase, hyaluronidase, and metalloproteinase activity of L. intermedia venom. This study suggests that this multiepitopic protein can be a suitable candidate for experimental vaccination approaches or for antivenom production against Loxosceles spp. venoms. PMID:29666624

Structural Basis for the Specific Neutralization of Stx2a with a Camelid Single Domain Antibody Fragment

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Robert Alvin Bernedo-Navarro

2018-03-01

Full Text Available Background: Shiga toxin-producing Escherichia coli (STEC are a subset of pathogens leading to illnesses such as diarrhea, hemolytic uremic syndrome and even death. The Shiga toxins are the main virulence factors and divided in two groups: Stx1 and Stx2, of which the latter is more frequently associated with severe pathologies in humans. Results: An immune library of nanobodies (Nbs was constructed after immunizing an alpaca with recombinant Shiga toxin-2a B subunit (rStx2aB, to retrieve multiple rStx2aB-specific Nbs. The specificity of five Nbs towards rStx2aB was confirmed in ELISA and Western blot. Nb113 had the highest affinity (9.6 nM and its bivalent construct exhibited a 100-fold higher functional affinity. The structure of the Nb113 in complex with rStx2aB was determined via X-ray crystallography. The crystal structure of the Nb113–rStx2aB complex revealed that five copies of Nb113 bind to the rStx2aB pentamer and that the Nb113 epitope overlaps with the Gb3 binding site, thereby providing a structural basis for the neutralization of Stx2a by Nb113 that was observed on Vero cells. Finally, the tandem-repeated, bivalent Nb1132 exhibits a higher toxin neutralization capacity compared to monovalent Nb113. Conclusions: The Nb of highest affinity for rStx2aB is also the best Stx2a and Stx2c toxin neutralizing Nb, especially in a bivalent format. This lead Nb neutralizes Stx2a by competing for the Gb3 receptor. The fusion of the bivalent Nb1132 with a serum albumin specific Nb is expected to combine high toxin neutralization potential with prolonged blood circulation.

Use of Monoclonal Antibodies in the Sensitive Detection and Neutralization of Botulinum Neurotoxin Serotype B

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Luisa W. Cheng

2015-11-01

Full Text Available Botulinum neurotoxins (BoNT are some of nature’s most potent toxins. Due to potential food contamination, and bioterrorism concerns, the development of detection reagents, therapeutics and countermeasures are of urgent interest. Recently, we have developed a sensitive electrochemiluminescent (ECL immunoassay for BoNT/B, using monoclonal antibodies (mAbs MCS6-27 and anti-BoNT/B rabbit polyclonal antibodies as the capture and detector. The ECL assay detected as little as 1 pg/mL BoNT/B in the buffer matrix, surpassing the detection sensitivities of the gold standard mouse bioassays. The ECL assay also allowed detection of BoNT/B in sera matrices of up to 100% sera with negligible matrix effects. This highly-sensitive assay allowed the determination of the biological half-lives of BoNT/B holotoxin in vivo. We further tested the toxin neutralization potential of our monoclonal antibodies using the mouse systemic and oral intoxication models. A combination of mAbs protected mice in both pre- and post-exposure models to lethal doses of BoNT/B. MAbs were capable of increasing survival of animals when administered even 10 h post-intoxication in an oral model, suggesting a likely time for BoNT/B complexes to reach the blood stream. More sensitive detection assays and treatments against BoNT intoxication will greatly enhance efforts to combat botulism.

MHC Class II and Non-MHC Class II Genes Differentially Influence Humoral Immunity to Bacillus anthracis Lethal Factor and Protective Antigen

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Judith A. James

2012-12-01

Full Text Available Anthrax Lethal Toxin consists of Protective Antigen (PA and Lethal Factor (LF, and current vaccination strategies focus on eliciting antibodies to PA. In human vaccination, the response to PA can vary greatly, and the response is often directed toward non-neutralizing epitopes. Variable vaccine responses have been shown to be due in part to genetic differences in individuals, with both MHC class II and other genes playing roles. Here, we investigated the relative contribution of MHC class II versus non-MHC class II genes in the humoral response to PA and LF immunization using three immunized strains of inbred mice: A/J (H-2k at the MHC class II locus, B6 (H-2b, and B6.H2k (H-2k. IgG antibody titers to LF were controlled primarily by the MHC class II locus, whereas IgG titers to PA were strongly influenced by the non-MHC class II genetic background. Conversely, the humoral fine specificity of reactivity to LF appeared to be controlled primarily through non-MHC class II genes, while the specificity of reactivity to PA was more dependent on MHC class II. Common epitopes, reactive in all strains, occurred in both LF and PA responses. These results demonstrate that MHC class II differentially influences humoral immune responses to LF and PA.

Staphylococcal Bicomponent Pore-Forming Toxins: Targets for Prophylaxis and Immunotherapy

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M. Javad Aman

2014-03-01

Full Text Available Staphylococccus aureus represents one of the most challenging human pathogens as well as a common colonizer of human skin and mucosal surfaces. S. aureus causes a wide range of diseases from skin and soft tissue infection (SSTI to debilitating and life-threatening conditions such as osteomyelitis, endocarditis, and necrotizing pneumonia. The range of diseases reflects the remarkable diversity of the virulence factors produced by this pathogen, including surface antigens involved in the establishment of infection and a large number of toxins that mediate a vast array of cellular responses. The staphylococcal toxins are generally believed to have evolved to disarm the innate immune system, the first line of defense against this pathogen. This review focuses on recent advances on elucidating the biological functions of S. aureus bicomponent pore-forming toxins (BCPFTs and their utility as targets for preventive and therapeutic intervention. These toxins are cytolytic to a variety of immune cells, primarily neutrophils, as well as cells with a critical barrier function. The lytic activity of BCPFTs towards immune cells implies a critical role in immune evasion, and a number of epidemiological studies and animal experiments relate these toxins to clinical disease, particularly SSTI and necrotizing pneumonia. Antibody-mediated neutralization of this lytic activity may provide a strategy for development of toxoid-based vaccines or immunotherapeutics for prevention or mitigation of clinical diseases. However, certain BCPFTs have been proposed to act as danger signals that may alert the immune system through an inflammatory response. The utility of a neutralizing vaccination strategy must be weighed against such immune-activating potential.

Staphylococcal bicomponent pore-forming toxins: targets for prophylaxis and immunotherapy.

Science.gov (United States)

Aman, M Javad; Adhikari, Rajan P

2014-03-04

Staphylococccus aureus represents one of the most challenging human pathogens as well as a common colonizer of human skin and mucosal surfaces. S. aureus causes a wide range of diseases from skin and soft tissue infection (SSTI) to debilitating and life-threatening conditions such as osteomyelitis, endocarditis, and necrotizing pneumonia. The range of diseases reflects the remarkable diversity of the virulence factors produced by this pathogen, including surface antigens involved in the establishment of infection and a large number of toxins that mediate a vast array of cellular responses. The staphylococcal toxins are generally believed to have evolved to disarm the innate immune system, the first line of defense against this pathogen. This review focuses on recent advances on elucidating the biological functions of S. aureus bicomponent pore-forming toxins (BCPFTs) and their utility as targets for preventive and therapeutic intervention. These toxins are cytolytic to a variety of immune cells, primarily neutrophils, as well as cells with a critical barrier function. The lytic activity of BCPFTs towards immune cells implies a critical role in immune evasion, and a number of epidemiological studies and animal experiments relate these toxins to clinical disease, particularly SSTI and necrotizing pneumonia. Antibody-mediated neutralization of this lytic activity may provide a strategy for development of toxoid-based vaccines or immunotherapeutics for prevention or mitigation of clinical diseases. However, certain BCPFTs have been proposed to act as danger signals that may alert the immune system through an inflammatory response. The utility of a neutralizing vaccination strategy must be weighed against such immune-activating potential.

Clostridium difficile 027/BI/NAP1 encodes a hypertoxic and antigenically variable form of TcdB.

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Jordi M Lanis

Full Text Available The Clostridium difficile exotoxin, TcdB, which is a major virulence factor, varies between strains of this pathogen. Herein, we show that TcdB from the epidemic BI/NAP1/027 strain of C. difficile is more lethal, causes more extensive brain hemorrhage, and is antigenically variable from TcdB produced by previously studied strains of this pathogen (TcdB003. In mouse intoxication assays, TcdB from a ribotype 027 strain (TcdB027 was at least four fold more lethal than TcdB003. TcdB027 caused a previously undescribed brain hemorrhage in mice and this correlated with a heightened sensitivity of brain microvascular endothelial cells to the toxin. TcdB003 and TcdB027 also differed in their antigenic profiles and did not share cross-neutralizing epitopes in a major immunogenic region of the protein. Solid phase humoral mapping of epitopes in the carboxy-terminal domains (CTD of TcdB027 and TcdB003 identified 11 reactive epitopes that varied between the two forms of TcdB, and 13 epitopes that were shared or overlapping. Despite the epitope differences and absence of neutralizing epitopes in the CTD of TcdB027, a toxoid form of this toxin primed a strong protective response. These findings indicate TcdB027 is a more potent toxin than TcdB003 as measured by lethality assays and pathology, moreover the sequence differences between the two forms of TcdB alter antigenic epitopes and reduce cross-neutralization by antibodies targeting the CTD.

Daboia russellii and Naja kaouthia venom neutralization by lupeol acetate isolated from the root extract of Indian sarsaparilla Hemidesmus indicus R.Br.

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Chatterjee, Ipshita; Chakravarty, A K; Gomes, A

2006-06-15

The present study reports the isolation and purification of lupeol acetate from the methanolic root extract of Indian medicinal plant Hemidesmus indicus (L.) R.Br. (family: Asclepiadaceae) which could neutralize venom induced action of Daboia russellii and Naja kaouthia on experimental animals. Lupeol acetate could significantly neutralize lethality, haemorrhage, defibrinogenation, edema, PLA(2) activity induced by Daboia russellii venom. It also neutralized Naja kaouthia venom induced lethality, cardiotoxicity, neurotoxicity and respiratory changes in experimental animals. Lupeol acetate potentiated the protection by snake venom antiserum action against Daboia russellii venom induced lethality in male albino mice. Venom induced changes in lipid peroxidation and super oxide dismutase activity was antagonized by lupeol acetate. Snake venom neutralization by lupeol acetate and its possible mechanism of action has been discussed.

Peptide Probes Reveal a Hydrophobic Steric Ratchet in the Anthrax Toxin Protective Antigen Translocase.

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Colby, Jennifer M; Krantz, Bryan A

2015-11-06

Anthrax toxin is a tripartite virulence factor produced by Bacillus anthracis during infection. Under acidic endosomal pH conditions, the toxin's protective antigen (PA) component forms a transmembrane channel in host cells. The PA channel then translocates its two enzyme components, lethal factor and edema factor, into the host cytosol under the proton motive force. Protein translocation under a proton motive force is catalyzed by a series of nonspecific polypeptide binding sites, called clamps. A 10-residue guest/host peptide model system, KKKKKXXSXX, was used to functionally probe polypeptide-clamp interactions within wild-type PA channels. The guest residues were Thr, Ala, Leu, Phe, Tyr, and Trp. In steady-state translocation experiments, the channel blocked most tightly with peptides that had increasing amounts of nonpolar surface area. Cooperative peptide binding was observed in the Trp-containing peptide sequence but not the other tested sequences. Trp substitutions into a flexible, uncharged linker between the lethal factor amino-terminal domain and diphtheria toxin A chain expedited translocation. Therefore, peptide-clamp sites in translocase channels can sense large steric features (like tryptophan) in peptides, and while these steric interactions may make a peptide translocate poorly, in the context of folded domains, they can make the protein translocate more rapidly presumably via a hydrophobic steric ratchet mechanism. Copyright © 2015 The Authors. Published by Elsevier Ltd.. All rights reserved.

The relative potency of inverse opioid agonists and a neutral opioid antagonist in precipitated withdrawal and antagonism of analgesia and toxicity.

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Sirohi, Sunil; Dighe, Shveta V; Madia, Priyanka A; Yoburn, Byron C

2009-08-01

Opioid antagonists can be classified as inverse agonists and neutral antagonists. In the opioid-dependent state, neutral antagonists are significantly less potent in precipitating withdrawal than inverse agonists. Consequently, neutral opioid antagonists may offer advantages over inverse agonists in the management of opioid overdose. In this study, the relative potency of three opioid antagonists to block opioid analgesia and toxicity and precipitate withdrawal was examined. First, the potency of two opioid inverse agonists (naltrexone and naloxone) and a neutral antagonist (6beta-naltrexol) to antagonize fentanyl-induced analgesia and lethality was determined. The order of potency to block analgesia was naltrexone > naloxone > 6beta-naltrexol (17, 4, 1), which was similar to that to block lethality (13, 2, 1). Next, the antagonists were compared using withdrawal jumping in fentanyl-dependent mice. The order of potency to precipitate withdrawal jumping was naltrexone > naloxone 6beta-naltrexol (1107, 415, 1). The relative potencies to precipitate withdrawal for the inverse agonists compared with the neutral antagonist were dramatically different from that for antagonism of analgesia and lethality. Finally, the effect of 6beta-naltrexol pretreatment on naloxone-precipitated jumping was determined in morphine and fentanyl-dependent mice. 6beta-Naltrexol pretreatment decreased naloxone precipitated withdrawal, indicating that 6beta-naltrexol is a neutral antagonist. These data demonstrate that inverse agonists and neutral antagonists have generally comparable potencies to block opioid analgesia and lethality, whereas the neutral opioid antagonist is substantially less potent in precipitating opioid withdrawal. These results support suggestions that neutral antagonists may have advantages over inverse agonists in the management of opioid overdose.

Noninvasive imaging technologies reveal edema toxin as a key virulence factor in anthrax.

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Dumetz, Fabien; Jouvion, Grégory; Khun, Huot; Glomski, Ian Justin; Corre, Jean-Philippe; Rougeaux, Clémence; Tang, Wei-Jen; Mock, Michèle; Huerre, Michel; Goossens, Pierre Louis

2011-06-01

Powerful noninvasive imaging technologies enable real-time tracking of pathogen-host interactions in vivo, giving access to previously elusive events. We visualized the interactions between wild-type Bacillus anthracis and its host during a spore infection through bioluminescence imaging coupled with histology. We show that edema toxin plays a central role in virulence in guinea pigs and during inhalational infection in mice. Edema toxin (ET), but not lethal toxin (LT), markedly modified the patterns of bacterial dissemination leading, to apparent direct dissemination to the spleen and provoking apoptosis of lymphoid cells. Each toxin alone provoked particular histological lesions in the spleen. When ET and LT are produced together during infection, a specific temporal pattern of lesion developed, with early lesions typical of LT, followed at a later stage by lesions typical of ET. Our study provides new insights into the complex spatial and temporal effects of B. anthracis toxins in the infected host, suggesting a greater role than previously suspected for ET in anthrax and suggesting that therapeutic targeting of ET contributes to protection. Copyright © 2011 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.

Efficacy of tannins from Mimosa pudica and tannic acid in neutralizing cobra (Naja kaouthia venom

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FY Sia

2011-01-01

Full Text Available In the present study, the effectiveness of Mimosa pudica tannins (MPT in neutralizing the lethality of Naja kaouthia venom was compared with commercially derived tannins. Preincubation of MPT with N. kaouthia venom maintained 100% survival of mice after 24 hours. The mouse group in which there was no preincubation, no protection against the effects of the venom was observed. M. pudica tannin was found to be more effective in neutralizing the lethality of N. kaouthia venom when compared to commercial tannic acid. Two protein spots were missing in the two-dimensional gel electrophoresis (2-DE of the MPT treated mouse indicating the down-regulation of venom proteins. The results from this study indicated that tannins obtained from M. pudica are better than tannic acid in neutralizing the lethality of N. kaouthia venom in vitro. However, further investigations are required to establish that M. pudica has potential for treating N. kaouthia snakebites.

Filaggrin-dependent secretion of sphingomyelinase protects against staphylococcal α-toxin-induced keratinocyte death.

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Brauweiler, Anne M; Bin, Lianghua; Kim, Byung Eui; Oyoshi, Michiko K; Geha, Raif S; Goleva, Elena; Leung, Donald Y M

2013-02-01

The skin of patients with atopic dermatitis (AD) has defects in keratinocyte differentiation, particularly in expression of the epidermal barrier protein filaggrin. AD skin lesions are often exacerbated by Staphylococcus aureus-mediated secretion of the virulence factor α-toxin. It is unknown whether lack of keratinocyte differentiation predisposes to enhanced lethality from staphylococcal toxins. We investigated whether keratinocyte differentiation and filaggrin expression protect against cell death induced by staphylococcal α-toxin. Filaggrin-deficient primary keratinocytes were generated through small interfering RNA gene knockdown. RNA expression was determined by using real-time PCR. Cell death was determined by using the lactate dehydrogenase assay. Keratinocyte cell survival in filaggrin-deficient (ft/ft) mouse skin biopsies was determined based on Keratin 5 staining. α-Toxin heptamer formation and acid sphingomyelinase expression were determined by means of immunoblotting. We found that filaggrin expression, occurring as the result of keratinocyte differentiation, significantly inhibits staphylococcal α-toxin-mediated pathogenicity. Furthermore, filaggrin plays a crucial role in protecting cells by mediating the secretion of sphingomyelinase, an enzyme that reduces the number of α-toxin binding sites on the keratinocyte surface. Finally, we determined that sphingomyelinase enzymatic activity directly prevents α-toxin binding and protects keratinocytes against α-toxin-induced cytotoxicity. The current study introduces the novel concept that S aureus α-toxin preferentially targets and destroys filaggrin-deficient keratinocytes. It also provides a mechanism to explain the increased propensity for S aureus-mediated exacerbation of AD skin disease. Copyright © 2012 American Academy of Allergy, Asthma & Immunology. Published by Mosby, Inc. All rights reserved.

LETHAL MUSHROOM TOXINS: ANALYSIS OF THE AMANITINS AND APPLICATION OF LATERAL FLOW IMMUNOASSAY

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KAYA, Ertuğrul

2018-01-01

Deaths from mushroom poisoning, due to the ingestionofmushrooms containing amatoxins, seem to be increasingboth in Turkey andworldwide. The amatoxinsfound in Amanita phalloides (also called thedeath capmushroom) are most toxic agents, and they are responsible for more than 95% ofthe cases of deadlymushroom poisoning. Alpha amanitin is best known toxin ofthis group. Alpha amanitin analysis can be carried out with special methodssuch as chromatography and ELISA. ELISA method is suitable in orde...

Importance of Neutralizing Monoclonal Antibodies Targeting Multiple Antigenic Sites on the Middle East Respiratory Syndrome Coronavirus Spike Glycoprotein To Avoid Neutralization Escape

Energy Technology Data Exchange (ETDEWEB)

Wang, Lingshu; Shi, Wei; Chappell, James D.; Joyce, M. Gordon; Zhang, Yi; Kanekiyo, Masaru; Becker, Michelle M.; van Doremalen, Neeltje; Fischer, Robert; Wang, Nianshuang; Corbett, Kizzmekia S.; Choe, Misook; Mason, Rosemarie D.; Van Galen, Joseph G.; Zhou, Tongqing; Saunders, Kevin O.; Tatti, Kathleen M.; Haynes, Lia M.; Kwong, Peter D.; Modjarrad, Kayvon; Kong, Wing-Pui; McLellan, Jason S.; Denison, Mark R.; Munster, Vincent J.; Mascola, John R.; Graham, Barney S.; Gallagher, Tom

2018-03-07

Middle East respiratory syndrome coronavirus (MERS-CoV) causes a highly lethal pulmonary infection with ~35% mortality. The potential for a future pandemic originating from animal reservoirs or health care-associated events is a major public health concern. There are no vaccines or therapeutic agents currently available for MERS-CoV. Using a probe-based single B cell cloning strategy, we have identified and characterized multiple neutralizing monoclonal antibodies (MAbs) specifically binding to the receptor-binding domain (RBD) or S1 (non-RBD) regions from a convalescent MERS-CoV-infected patient and from immunized rhesus macaques. RBD-specific MAbs tended to have greater neutralizing potency than non-RBD S1-specific MAbs. Six RBD-specific and five S1-specific MAbs could be sorted into four RBD and three non-RBD distinct binding patterns, based on competition assays, mapping neutralization escape variants, and structural analysis. We determined cocrystal structures for two MAbs targeting the RBD from different angles and show they can bind the RBD only in the “out” position. We then showed that selected RBD-specific, non-RBD S1-specific, and S2-specific MAbs given prophylactically prevented MERS-CoV replication in lungs and protected mice from lethal challenge. Importantly, combining RBD- and non-RBD MAbs delayed the emergence of escape mutations in a cell-based virus escape assay. These studies identify MAbs targeting different antigenic sites on S that will be useful for defining mechanisms of MERS-CoV neutralization and for developing more effective interventions to prevent or treat MERS-CoV infections.

IMPORTANCEMERS-CoV causes a highly lethal respiratory infection for which no vaccines or antiviral therapeutic options are currently available. Based on continuing exposure from established reservoirs in dromedary camels and bats, transmission of MERS-CoV into humans and future outbreaks are expected. Using

A Medical Research and Evaluation Facility (MREF) and Studies Supporting the Medical Chemical Defense Program. Evaluation of the Passive Protection Against Five Serotypes of Botulinum Toxin Provided by Botulinum Human Immune Globulin in an Animal Model

National Research Council Canada - National Science Library

Olson, Carl

1998-01-01

Pentavalent (ABCDE) botulinum toxoid vaccine is intended for use as a prophylactic measure to protect combat troops against the lethal effects of botulinum toxins A-E, a group of toxins considered to be a serious biological warfare threat...

The scorpion toxin Bot IX is a potent member of the α-like family and has a unique N-terminal sequence extension.

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Martin-Eauclaire, Marie-France; Salvatierra, Juan; Bosmans, Frank; Bougis, Pierre E

2016-09-01

We report the detailed chemical, immunological and pharmacological characterization of the α-toxin Bot IX from the Moroccan scorpion Buthus occitanus tunetanus venom. Bot IX, which consists of 70 amino acids, is a highly atypical toxin. It carries a unique N-terminal sequence extension and is highly lethal in mice. Voltage clamp recordings on oocytes expressing rat Nav1.2 or insect BgNav1 reveal that, similar to other α-like toxins, Bot IX inhibits fast inactivation of both variants. Moreover, Bot IX belongs to the same structural/immunological group as the α-like toxin Bot I. Remarkably, radioiodinated Bot IX competes efficiently with the classical α-toxin AaH II from Androctonus australis, and displays one of the highest affinities for Nav channels. © 2016 Federation of European Biochemical Societies.

Protective Monotherapy Against Lethal Ebola Virus Infection by a Potently Neutralizing Antibody

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2016-07-11

were 49   identified and enrolled in VRC200 clinical trial #NCT00067054 after giving signed 50   informed consent . Peripheral blood mononuclear...illness 56   when administered one day after lethal challenge. Treatment with a single human 57   mAb suggests a simplified therapeutic strategy for...efforts to simplify the ZMapp regimen to contain fewer mAbs have not been successful in 75   the macaque EVD model (7). We sought to isolate

Targeting Staphylococcus aureus Toxins: A Potential form of Anti-Virulence Therapy

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Cin Kong

2016-03-01

Full Text Available Staphylococcus aureus is an opportunistic pathogen and the leading cause of a wide range of severe clinical infections. The range of diseases reflects the diversity of virulence factors produced by this pathogen. To establish an infection in the host, S. aureus expresses an inclusive set of virulence factors such as toxins, enzymes, adhesins, and other surface proteins that allow the pathogen to survive under extreme conditions and are essential for the bacteria’s ability to spread through tissues. Expression and secretion of this array of toxins and enzymes are tightly controlled by a number of regulatory systems. S. aureus is also notorious for its ability to resist the arsenal of currently available antibiotics and dissemination of various multidrug-resistant S. aureus clones limits therapeutic options for a S. aureus infection. Recently, the development of anti-virulence therapeutics that neutralize S. aureus toxins or block the pathways that regulate toxin production has shown potential in thwarting the bacteria’s acquisition of antibiotic resistance. In this review, we provide insights into the regulation of S. aureus toxin production and potential anti-virulence strategies that target S. aureus toxins.

Therapeutic Approaches of Botulinum Toxin in Gynecology.

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Moga, Marius Alexandru; Dimienescu, Oana Gabriela; Bălan, Andreea; Scârneciu, Ioan; Barabaș, Barna; Pleș, Liana

2018-04-21

Botulinum toxins (BoNTs) are produced by several anaerobic species of the genus Clostridium and, although they were originally considered lethal toxins, today they find their usefulness in the treatment of a wide range of pathologies in various medical specialties. Botulinum neurotoxin has been identified in seven different isoforms (BoNT-A, BoNT-B, BoNT-C, BoNT-D, BoNT-E, BoNT-F, and BoNT-G). Neurotoxigenic Clostridia can produce more than 40 different BoNT subtypes and, recently, a new BoNT serotype (BoNT-X) has been reported in some studies. BoNT-X has not been shown to actually be an active neurotoxin despite its catalytically active LC, so it should be described as a putative eighth serotype. The mechanism of action of the serotypes is similar: they inhibit the release of acetylcholine from the nerve endings but their therapeutically potency varies. Botulinum toxin type A (BoNT-A) is the most studied serotype for therapeutic purposes. Regarding the gynecological pathology, a series of studies based on the efficiency of its use in the treatment of refractory myofascial pelvic pain, vaginism, dyspareunia, vulvodynia and overactive bladder or urinary incontinence have been reported. The current study is a review of the literature regarding the efficiency of BoNT-A in the gynecological pathology and on the long and short-term effects of its administration.

Glycoprotein-Specific Antibodies Produced by DNA Vaccination Protect Guinea Pigs from Lethal Argentine and Venezuelan Hemorrhagic Fever

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Golden, Joseph W.; Maes, Piet; Kwilas, Steven A.; Ballantyne, John

2016-01-01

ABSTRACT Several members of the Arenaviridae can cause acute febrile diseases in humans, often resulting in lethality. The use of convalescent-phase human plasma is an effective treatment in humans infected with arenaviruses, particularly species found in South America. Despite this, little work has focused on developing potent and defined immunotherapeutics against arenaviruses. In the present study, we produced arenavirus neutralizing antibodies by DNA vaccination of rabbits with plasmids encoding the full-length glycoprotein precursors of Junín virus (JUNV), Machupo virus (MACV), and Guanarito virus (GTOV). Geometric mean neutralizing antibody titers, as measured by the 50% plaque reduction neutralization test (PRNT50), exceeded 5,000 against homologous viruses. Antisera against each targeted virus exhibited limited cross-species binding and, to a lesser extent, cross-neutralization. Anti-JUNV glycoprotein rabbit antiserum protected Hartley guinea pigs from lethal intraperitoneal infection with JUNV strain Romero when the antiserum was administered 2 days after challenge and provided some protection (∼30%) when administered 4 days after challenge. Treatment starting on day 6 did not protect animals. We further formulated an IgG antibody cocktail by combining anti-JUNV, -MACV, and -GTOV antibodies produced in DNA-vaccinated rabbits. This cocktail protected 100% of guinea pigs against JUNV and GTOV lethal disease. We then expanded on this cocktail approach by simultaneously vaccinating rabbits with a combination of plasmids encoding glycoproteins from JUNV, MACV, GTOV, and Sabia virus (SABV). Sera collected from rabbits vaccinated with the combination vaccine neutralized all four targets. These findings support the concept of using a DNA vaccine approach to generate a potent pan-arenavirus immunotherapeutic. IMPORTANCE Arenaviruses are an important family of emerging viruses. In infected humans, convalescent-phase plasma containing neutralizing antibodies can

Variability in mutational fitness effects prevents full lethal transitions in large quasispecies populations

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Sardanyés, Josep; Simó, Carles; Martínez, Regina; Solé, Ricard V.; Elena, Santiago F.

2014-04-01

The distribution of mutational fitness effects (DMFE) is crucial to the evolutionary fate of quasispecies. In this article we analyze the effect of the DMFE on the dynamics of a large quasispecies by means of a phenotypic version of the classic Eigen's model that incorporates beneficial, neutral, deleterious, and lethal mutations. By parameterizing the model with available experimental data on the DMFE of Vesicular stomatitis virus (VSV) and Tobacco etch virus (TEV), we found that increasing mutation does not totally push the entire viral quasispecies towards deleterious or lethal regions of the phenotypic sequence space. The probability of finding regions in the parameter space of the general model that results in a quasispecies only composed by lethal phenotypes is extremely small at equilibrium and in transient times. The implications of our findings can be extended to other scenarios, such as lethal mutagenesis or genomically unstable cancer, where increased mutagenesis has been suggested as a potential therapy.

Rabbit IgG antibodies against Phospholipase A2 from Crotalus durissus terrificus neutralize the lethal activity of the venom Los anticuerpos IgG de conejos anti-fosfolipasa A2 de Crotalus durissus terrificus neutralizan la actividad letal del veneno

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Juan P. Rodríguez

2006-12-01

Full Text Available Crotalus durissus terrificus (C.d.t. (South American rattlesnake venom possesses myotoxic and neurotoxic activities, both of which are also expressed by crotoxin, the principal toxin of this venom. Crotoxin contains a basic phospholipase A2 (PLA2 and a non toxic acidic protein, crotapotin. We have produced and investigated the ability of IgG antibodies raised in rabbits against PLA2 to neutralize the lethality of the whole venom. PLA2 was isolated by gel filtration chromatography (Sephadex G-75. Specific antibodies were obtained by subcutaneous and intramuscular inoculation of PLA2 (700 µg with Freund adjuvant. Groups of six mice (20 + 2 g were inoculated with 0.5 ml i.p. of C. d. t. venom (4 µg or a mixture of venom that had been preincubated with the desired volume of IgG antibodies. Mortality, recorded 24 and 48 h after inoculation, showed that IgG anti-PLA2 were more effective than anticrotalic serum in neutralizing the lethal activity. These results demonstrate that it could be possible to obtain an anti-venom made by specific antibodies with a high level of protection against the lethal component of C.d.t. venom, and/or the inclusion of these antibodies as a supplement in heterologous anti-venoms.El veneno de Crotalus durissus terrificus (C.d.t. (Cascabel de Sud América posee actividad miotóxica y neurotóxica, actividades que también exhibe el complejo crotoxina, principal componente tóxico de este veneno. El complejo crotoxina está constituido por una fosfolipasa A2 básica (PLA2 y una proteína acídica no tóxica, el crotapotín. En este trabajo se estudió la capacidad neutralizante de anticuerpos IgG anti-PLA2 sobre la letalidad inducida por el veneno entero. El antígeno PLA2, fue aislado por cromatografía de filtración en gel (Sephadex G-75. Se inocularon conejos machos por vía subcutánea e intramuscular, con 700 µg de PLA2 y adyuvante para la obtención de anticuerpos específicos. La capacidad neutralizante del

The Tip of the Four N-Terminal α-Helices of Clostridium sordellii Lethal Toxin Contains the Interaction Site with Membrane Phosphatidylserine Facilitating Small GTPases Glucosylation

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Carolina Varela Chavez

2016-03-01

Full Text Available Clostridium sordellii lethal toxin (TcsL is a powerful virulence factor responsible for severe toxic shock in man and animals. TcsL belongs to the large clostridial glucosylating toxin (LCGT family which inactivates small GTPases by glucosylation with uridine-diphosphate (UDP-glucose as a cofactor. Notably, TcsL modifies Rac and Ras GTPases, leading to drastic alteration of the actin cytoskeleton and cell viability. TcsL enters cells via receptor-mediated endocytosis and delivers the N-terminal glucosylating domain (TcsL-cat into the cytosol. TcsL-cat was found to preferentially bind to phosphatidylserine (PS-containing membranes and to increase the glucosylation of Rac anchored to the lipid membrane. We have previously reported that the N-terminal four helical bundle structure (1–93 domain recognizes a broad range of lipids, but that TcsL-cat specifically binds to PS and phosphatidic acid. Here, we show using mutagenesis that the PS binding site is localized on the tip of the four-helix bundle which is rich in positively-charged amino acids. Residues Y14, V15, F17, and R18 on loop 1, between helices 1 and 2, in coordination with R68 from loop 3, between helices 3 and 4, form a pocket which accommodates L-serine. The functional PS-binding site is required for TcsL-cat binding to the plasma membrane and subsequent cytotoxicity. TcsL-cat binding to PS facilitates a high enzymatic activity towards membrane-anchored Ras by about three orders of magnitude as compared to Ras in solution. The PS-binding site is conserved in LCGTs, which likely retain a common mechanism of binding to the membrane for their full activity towards membrane-bound GTPases.

Identification and characterization of two novel toxins expressed by the lethal honey bee pathogen Paenibacillus larvae, the causative agent of American foulbrood.

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Fünfhaus, Anne; Poppinga, Lena; Genersch, Elke

2013-11-01

Paenibacillus larvae is a Gram-positive bacterial pathogen causing the epizootic American foulbrood in honey bee larvae. Four so-called enterobacterial repetitive intergenic consensus (ERIC) genotypes of P. larvae exist with P. larvae genotypes ERIC I and ERIC II being responsible for disease outbreaks all over the world. Very few molecular data on the pathogen, on pathogenesis or on virulence factors exist. We now identified two genomic loci in P. larvae ERIC I coding for two binary AB toxins, Plx1 and Plx2. In silico analyses revealed that Plx1 is the third member of an enigmatic family of AB toxins so far only comprising MTX1 of Lysinibacillus sphaericus and pierisin-like toxins expressed by several butterflies. Plx2 is also remarkable because the A-domain is highly similar to C3 exoenzymes, which normally are single domain proteins, while the B-domain is homologous to B-domains of C2-toxins. We constructed P. larvae mutants lacking expression of Plx1, Plx2 or both toxins and demonstrated that these toxins are important virulence factors for P. larvae ERIC I. © 2013 Society for Applied Microbiology and John Wiley & Sons Ltd.

Quinoid radio-toxin (QRT) induced metabolic changes in mice: An ex vivo and in vivo EPR investigation

Science.gov (United States)

Ibragimova, M.I.; Petukhov, V.Yu.; Zheglov, E.P.; Khan, N.; Hou, H.; Swartz, H.M.; Konjukhov, G.V.; Nizamov, R.N.

2013-01-01

Radio-toxins are toxic metabolites produced by ionizing irradiation and have toxic effects similar to those caused by direct irradiation. We have investigated the effect of a quinoid radio-toxin (QRT) obtained from γ-irradiated potato tuber on various organs in mice using ex vivo and in vivo EPR spectroscopy. Results indicate a decrease in the activity of ribonucleotide reductase enzyme in spleen of mice treated with 0.2 mg QRT. A dose of 2 mg QRT was fatal to mice within 45–60 min of treatment. Nitrosyl hemoglobin complexes α-(Fe2+–NO)α-(Fe2+)β-(Fe2+)2 were detected from spleen, blood, liver, kidney, heart, and lung tissue samples of mice treated with lethal doses of QRT. A significant decrease of pO2 in liver and brain was observed after administration of QRT at the lethal dose. The time of the appearance of the nitrosyl hemoglobin complex and its intensity varied with the dose of QRT and the type of tissue. These results indicate that the effect of the QRT is more prominent in spleen and to a lesser extent in liver and blood. The QRT action at the lethal doses resulted in an increased hypoxia over time with disruption of compensatory adaptive response. The results indicate similar outcome of QRT as observed with γ-irradiation. PMID:18230367

Antibody-Mediated Neutralization of the Exotoxin Mycolactone, the Main Virulence Factor Produced by Mycobacterium ulcerans.

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Jean-Pierre Dangy

2016-06-01

Full Text Available Mycolactone, the macrolide exotoxin produced by Mycobacterium ulcerans, causes extensive tissue destruction by inducing apoptosis of host cells. In this study, we aimed at the production of antibodies that could neutralize the cytotoxic activities of mycolactone.Using the B cell hybridoma technology, we generated a series of monoclonal antibodies with specificity for mycolactone from spleen cells of mice immunized with the protein conjugate of a truncated synthetic mycolactone derivative. L929 fibroblasts were used as a model system to investigate whether these antibodies can inhibit the biological effects of mycolactone. By measuring the metabolic activity of the fibroblasts, we found that anti-mycolactone mAbs can completely neutralize the cytotoxic activity of mycolactone.The toxin neutralizing capacity of anti-mycolactone mAbs supports the concept of evaluating the macrolide toxin as vaccine target.

Ophiophagus hannah Venom: Proteome, Components Bound by Naja kaouthia Antivenin and Neutralization by N. kaouthia Neurotoxin-Specific Human ScFv

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Witchuda Danpaiboon

2014-05-01

Full Text Available Venomous snakebites are an important health problem in tropical and subtropical countries. King cobra (Ophiophagus hannah is the largest venomous snake found in South and Southeast Asia. In this study, the O. hannah venom proteome and the venom components cross-reactive to N. kaouthia monospecific antivenin were studied. O. hannah venom consisted of 14 different protein families, including three finger toxins, phospholipases, cysteine-rich secretory proteins, cobra venom factor, muscarinic toxin, L-amino acid oxidase, hypothetical proteins, low cysteine protein, phosphodiesterase, proteases, vespryn toxin, Kunitz, growth factor activators and others (coagulation factor, endonuclease, 5’-nucleotidase. N. kaouthia antivenin recognized several functionally different O. hannah venom proteins and mediated paratherapeutic efficacy by rescuing the O. hannah envenomed mice from lethality. An engineered human ScFv specific to N. kaouthia long neurotoxin (NkLN-HuScFv cross-neutralized the O. hannah venom and extricated the O. hannah envenomed mice from death in a dose escalation manner. Homology modeling and molecular docking revealed that NkLN-HuScFv interacted with residues in loops 2 and 3 of the neurotoxins of both snake species, which are important for neuronal acetylcholine receptor binding. The data of this study are useful for snakebite treatment when and where the polyspecific antivenin is not available. Because the supply of horse-derived antivenin is limited and the preparation may cause some adverse effects in recipients, a cocktail of recombinant human ScFvs for various toxic venom components shared by different venomous snakes, exemplified by the in vitro produced NkLN-HuScFv in this study, should contribute to a possible future route for an improved alternative to the antivenins.

Ophiophagus hannah venom: proteome, components bound by Naja kaouthia antivenin and neutralization by N. kaouthia neurotoxin-specific human ScFv.

Science.gov (United States)

Danpaiboon, Witchuda; Reamtong, Onrapak; Sookrung, Nitat; Seesuay, Watee; Sakolvaree, Yuwaporn; Thanongsaksrikul, Jeeraphong; Dong-din-on, Fonthip; Srimanote, Potjanee; Thueng-in, Kanyarat; Chaicumpa, Wanpen

2014-05-13

Venomous snakebites are an important health problem in tropical and subtropical countries. King cobra (Ophiophagus hannah) is the largest venomous snake found in South and Southeast Asia. In this study, the O. hannah venom proteome and the venom components cross-reactive to N. kaouthia monospecific antivenin were studied. O. hannah venom consisted of 14 different protein families, including three finger toxins, phospholipases, cysteine-rich secretory proteins, cobra venom factor, muscarinic toxin, L-amino acid oxidase, hypothetical proteins, low cysteine protein, phosphodiesterase, proteases, vespryn toxin, Kunitz, growth factor activators and others (coagulation factor, endonuclease, 5'-nucleotidase). N. kaouthia antivenin recognized several functionally different O. hannah venom proteins and mediated paratherapeutic efficacy by rescuing the O. hannah envenomed mice from lethality. An engineered human ScFv specific to N. kaouthia long neurotoxin (NkLN-HuScFv) cross-neutralized the O. hannah venom and extricated the O. hannah envenomed mice from death in a dose escalation manner. Homology modeling and molecular docking revealed that NkLN-HuScFv interacted with residues in loops 2 and 3 of the neurotoxins of both snake species, which are important for neuronal acetylcholine receptor binding. The data of this study are useful for snakebite treatment when and where the polyspecific antivenin is not available. Because the supply of horse-derived antivenin is limited and the preparation may cause some adverse effects in recipients, a cocktail of recombinant human ScFvs for various toxic venom components shared by different venomous snakes, exemplified by the in vitro produced NkLN-HuScFv in this study, should contribute to a possible future route for an improved alternative to the antivenins.

Glycoprotein-Specific Antibodies Produced by DNA Vaccination Protect Guinea Pigs from Lethal Argentine and Venezuelan Hemorrhagic Fever.

Science.gov (United States)

Golden, Joseph W; Maes, Piet; Kwilas, Steven A; Ballantyne, John; Hooper, Jay W

2016-01-20

Several members of the Arenaviridae can cause acute febrile diseases in humans, often resulting in lethality. The use of convalescent-phase human plasma is an effective treatment in humans infected with arenaviruses, particularly species found in South America. Despite this, little work has focused on developing potent and defined immunotherapeutics against arenaviruses. In the present study, we produced arenavirus neutralizing antibodies by DNA vaccination of rabbits with plasmids encoding the full-length glycoprotein precursors of Junín virus (JUNV), Machupo virus (MACV), and Guanarito virus (GTOV). Geometric mean neutralizing antibody titers, as measured by the 50% plaque reduction neutralization test (PRNT(50)), exceeded 5,000 against homologous viruses. Antisera against each targeted virus exhibited limited cross-species binding and, to a lesser extent, cross-neutralization. Anti-JUNV glycoprotein rabbit antiserum protected Hartley guinea pigs from lethal intraperitoneal infection with JUNV strain Romero when the antiserum was administered 2 days after challenge and provided some protection (∼30%) when administered 4 days after challenge. Treatment starting on day 6 did not protect animals. We further formulated an IgG antibody cocktail by combining anti-JUNV, -MACV, and -GTOV antibodies produced in DNA-vaccinated rabbits. This cocktail protected 100% of guinea pigs against JUNV and GTOV lethal disease. We then expanded on this cocktail approach by simultaneously vaccinating rabbits with a combination of plasmids encoding glycoproteins from JUNV, MACV, GTOV, and Sabia virus (SABV). Sera collected from rabbits vaccinated with the combination vaccine neutralized all four targets. These findings support the concept of using a DNA vaccine approach to generate a potent pan-arenavirus immunotherapeutic. Arenaviruses are an important family of emerging viruses. In infected humans, convalescent-phase plasma containing neutralizing antibodies can mitigate the

Therapeutic Approaches of Botulinum Toxin in Gynecology

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Marius Alexandru Moga

2018-04-01

Full Text Available Botulinum toxins (BoNTs are produced by several anaerobic species of the genus Clostridium and, although they were originally considered lethal toxins, today they find their usefulness in the treatment of a wide range of pathologies in various medical specialties. Botulinum neurotoxin has been identified in seven different isoforms (BoNT-A, BoNT-B, BoNT-C, BoNT-D, BoNT-E, BoNT-F, and BoNT-G. Neurotoxigenic Clostridia can produce more than 40 different BoNT subtypes and, recently, a new BoNT serotype (BoNT-X has been reported in some studies. BoNT-X has not been shown to actually be an active neurotoxin despite its catalytically active LC, so it should be described as a putative eighth serotype. The mechanism of action of the serotypes is similar: they inhibit the release of acetylcholine from the nerve endings but their therapeutically potency varies. Botulinum toxin type A (BoNT-A is the most studied serotype for therapeutic purposes. Regarding the gynecological pathology, a series of studies based on the efficiency of its use in the treatment of refractory myofascial pelvic pain, vaginism, dyspareunia, vulvodynia and overactive bladder or urinary incontinence have been reported. The current study is a review of the literature regarding the efficiency of BoNT-A in the gynecological pathology and on the long and short-term effects of its administration.

Snake venomics of Crotalus tigris: the minimalist toxin arsenal of the deadliest Nearctic rattlesnake venom. Evolutionary Clues for generating a pan-specific antivenom against crotalid type II venoms [corrected].

Science.gov (United States)

Calvete, Juan J; Pérez, Alicia; Lomonte, Bruno; Sánchez, Elda E; Sanz, Libia

2012-02-03

We report the proteomic and antivenomic characterization of Crotalus tigris venom. This venom exhibits the highest lethality for mice among rattlesnakes and the simplest toxin proteome reported to date. The venom proteome of C. tigris comprises 7-8 gene products from 6 toxin families; the presynaptic β-neurotoxic heterodimeric PLA(2), Mojave toxin, and two serine proteinases comprise, respectively, 66 and 27% of the C. tigris toxin arsenal, whereas a VEGF-like protein, a CRISP molecule, a medium-sized disintegrin, and 1-2 PIII-SVMPs each represent 0.1-5% of the total venom proteome. This toxin profile really explains the systemic neuro- and myotoxic effects observed in envenomated animals. In addition, we found that venom lethality of C. tigris and other North American rattlesnake type II venoms correlates with the concentration of Mojave toxin A-subunit, supporting the view that the neurotoxic venom phenotype of crotalid type II venoms may be described as a single-allele adaptation. Our data suggest that the evolutionary trend toward neurotoxicity, which has been also reported for the South American rattlesnakes, may have resulted by pedomorphism. The ability of an experimental antivenom to effectively immunodeplete proteins from the type II venoms of C. tigris, Crotalus horridus , Crotalus oreganus helleri, Crotalus scutulatus scutulatus, and Sistrurus catenatus catenatus indicated the feasibility of generating a pan-American anti-Crotalus type II antivenom, suggested by the identification of shared evolutionary trends among South and North American Crotalus species.

Collaborative study for establishment of the European Pharmacopoeia BRP batch 1 for diphtheria toxin.

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Sesardic, D; Prior, C; Daas, A; Buchheit, K H

2003-07-01

A stable liquid candidate Biological Reference Preparation (BRP) for diphtheria toxin was prepared in peptone buffer (nominal content of diphtheria toxin: 1 Lf/ml, 0.4 micro g/ml), filled in ampoules (filling volume: 1 ml) and characterised in a collaborative study. The toxin is to be used in the test "Absence of toxin and irreversibility of toxoid" as described in the current European Pharmacopoeia (Ph. Eur.) monograph Diphtheria Vaccine (Adsorbed) (2002:0443). Eleven laboratories assessed the specific activity of the preparation by in vivo and in vitro assays. The material is assumed to have satisfactory stability with a calculated predicted loss of activity of LD( 50)/ml (lethal challenge) and >75 000 Lr/Lf (intradermal challenge). The candidate BRP was successfully used in nine laboratories and confirmed suitable for use in the Vero cell test for "Absence of toxin and irreversibility of toxoid" as described in the Ph. Eur. monograph 2002:0443; i.e., concentrations of 5 x 10( -5) Lf/ml and below caused cytotoxic effects in the Vero cell test. Due to its liquid nature, the stability of the material will be monitored at regular intervals and preparation of a stable freeze-dried formulation will be considered for long-term use. Additional studies will be performed to confirm suitability of this BRP for other applications. The candidate BRP was adopted as the Ph. Eur. reference material for Diphtheria Toxin Batch 1 by the Ph. Eur. Commission at its session in March 2003.

Viper and cobra venom neutralization by beta-sitosterol and stigmasterol isolated from the root extract of Pluchea indica Less. (Asteraceae).

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Gomes, A; Saha, Archita; Chatterjee, Ipshita; Chakravarty, A K

2007-09-01

We reported previously that the methanolic root extract of the Indian medicinal plant Pluchea indica Less. (Asteraceae) could neutralize viper venom-induced action [Alam, M.I., Auddy, B., Gomes, A., 1996. Viper venom neutralization by Indian medicinal plant (Hemidesmus indicus and P. indica) root extracts. Phytother. Res. 10, 58-61]. The present study reports the neutralization of viper and cobra venom by beta-sitosterol and stigmasterol isolated from the root extract of P. indica Less. (Asteraceae). The active fraction (containing the major compound beta-sitosterol and the minor compound stigmasterol) was isolated and purified by silica gel column chromatography and the structure was determined using spectroscopic analysis (EIMS, (1)H NMR, (13)C NMR). Anti-snake venom activity was studied in experimental animals. The active fraction was found to significantly neutralize viper venom-induced lethal, hemorrhagic, defibrinogenation, edema and PLA(2) activity. Cobra venom-induced lethality, cardiotoxicity, neurotoxicity, respiratory changes and PLA(2) activity were also antagonized by the active component. It potentiated commercial snake venom antiserum action against venom-induced lethality in male albino mice. The active fraction could antagonize venom-induced changes in lipid peroxidation and superoxide dismutase activity. This study suggests that beta-sitosterol and stigmasterol may play an important role, along with antiserum, in neutralizing snake venom-induced actions.

Potency of a human monoclonal antibody to diphtheria toxin relative to equine diphtheria anti-toxin in a guinea pig intoxication model.

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Smith, Heidi L; Cheslock, Peter; Leney, Mark; Barton, Bruce; Molrine, Deborah C

2016-08-17

Prompt administration of anti-toxin reduces mortality following Corynebacterium diphtheriae infection. Current treatment relies upon equine diphtheria anti-toxin (DAT), with a 10% risk of serum sickness and rarely anaphylaxis. The global DAT supply is extremely limited; most manufacturers have ceased production. S315 is a neutralizing human IgG1 monoclonal antibody to diphtheria toxin that may provide a safe and effective alternative to equine DAT and address critical supply issues. To guide dose selection for IND-enabling pharmacology and toxicology studies, we dose-ranged S315 and DAT in a guinea pig model of diphtheria intoxication based on the NIH Minimum Requirements potency assay. Animals received a single injection of antibody premixed with toxin, were monitored for 30 days, and assigned a numeric score for clinical signs of disease. Animals receiving ≥ 27.5 µg of S315 or ≥ 1.75 IU of DAT survived whereas animals receiving ≤ 22.5 µg of S315 or ≤ 1.25 IU of DAT died, yielding a potency estimate of 17 µg S315/IU DAT (95% CI 16-21) for an endpoint of survival. Because some surviving animals exhibited transient limb weakness, likely a systemic sign of toxicity, DAT and S315 doses required to prevent hind limb paralysis were also determined, yielding a relative potency of 48 µg/IU (95% CI 38-59) for this alternate endpoint. To support advancement of S315 into clinical trials, potency estimates will be used to evaluate the efficacy of S315 versus DAT in an animal model with antibody administration after toxin exposure, more closely modeling anti-toxin therapy in humans.

Ebolavirus Glycoprotein Fc Fusion Protein Protects Guinea Pigs against Lethal Challenge

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Konduru, Krishnamurthy; Shurtleff, Amy C.; Bradfute, Steven B.; Nakamura, Siham; Bavari, Sina; Kaplan, Gerardo

2016-01-01

Ebola virus (EBOV), a member of the Filoviridae that can cause severe hemorrhagic fever in humans and nonhuman primates, poses a significant threat to the public health. Currently, there are no licensed vaccines or therapeutics to prevent and treat EBOV infection. Several vaccines based on the EBOV glycoprotein (GP) are under development, including vectored, virus-like particles, and protein-based subunit vaccines. We previously demonstrated that a subunit vaccine containing the extracellular domain of the Ebola ebolavirus (EBOV) GP fused to the Fc fragment of human IgG1 (EBOVgp-Fc) protected mice against EBOV lethal challenge. Here, we show that the EBOVgp-Fc vaccine formulated with QS-21, alum, or polyinosinic-polycytidylic acid-poly-L-lysine carboxymethylcellulose (poly-ICLC) adjuvants induced strong humoral immune responses in guinea pigs. The vaccinated animals developed anti-GP total antibody titers of approximately 105−106 and neutralizing antibody titers of approximately 103 as assessed by a BSL-2 neutralization assay based on vesicular stomatitis virus (VSV) pseudotypes. The poly-ICLC formulated EBOVgp-Fc vaccine protected all the guinea pigs against EBOV lethal challenge performed under BSL-4 conditions whereas the same vaccine formulated with QS-21 or alum only induced partial protection. Vaccination with a mucin-deleted EBOVgp-Fc construct formulated with QS-21 adjuvant did not have a significant effect in anti-GP antibody levels and protection against EBOV lethal challenge compared to the full-length GP construct. The bulk of the humoral response induced by the EBOVgp-Fc vaccine was directed against epitopes outside the EBOV mucin region. Our findings indicate that different adjuvants can eliciting varying levels of protection against lethal EBOV challenge in guinea pigs vaccinated with EBOVgp-Fc, and suggest that levels of total anti-GP antibodies elicit by protein-based GP subunit vaccines do not correlate with protection. Our data further support

Ebolavirus Glycoprotein Fc Fusion Protein Protects Guinea Pigs against Lethal Challenge.

Science.gov (United States)

Konduru, Krishnamurthy; Shurtleff, Amy C; Bradfute, Steven B; Nakamura, Siham; Bavari, Sina; Kaplan, Gerardo

2016-01-01

Ebola virus (EBOV), a member of the Filoviridae that can cause severe hemorrhagic fever in humans and nonhuman primates, poses a significant threat to the public health. Currently, there are no licensed vaccines or therapeutics to prevent and treat EBOV infection. Several vaccines based on the EBOV glycoprotein (GP) are under development, including vectored, virus-like particles, and protein-based subunit vaccines. We previously demonstrated that a subunit vaccine containing the extracellular domain of the Ebola ebolavirus (EBOV) GP fused to the Fc fragment of human IgG1 (EBOVgp-Fc) protected mice against EBOV lethal challenge. Here, we show that the EBOVgp-Fc vaccine formulated with QS-21, alum, or polyinosinic-polycytidylic acid-poly-L-lysine carboxymethylcellulose (poly-ICLC) adjuvants induced strong humoral immune responses in guinea pigs. The vaccinated animals developed anti-GP total antibody titers of approximately 105-106 and neutralizing antibody titers of approximately 103 as assessed by a BSL-2 neutralization assay based on vesicular stomatitis virus (VSV) pseudotypes. The poly-ICLC formulated EBOVgp-Fc vaccine protected all the guinea pigs against EBOV lethal challenge performed under BSL-4 conditions whereas the same vaccine formulated with QS-21 or alum only induced partial protection. Vaccination with a mucin-deleted EBOVgp-Fc construct formulated with QS-21 adjuvant did not have a significant effect in anti-GP antibody levels and protection against EBOV lethal challenge compared to the full-length GP construct. The bulk of the humoral response induced by the EBOVgp-Fc vaccine was directed against epitopes outside the EBOV mucin region. Our findings indicate that different adjuvants can eliciting varying levels of protection against lethal EBOV challenge in guinea pigs vaccinated with EBOVgp-Fc, and suggest that levels of total anti-GP antibodies elicit by protein-based GP subunit vaccines do not correlate with protection. Our data further support

Ebolavirus Glycoprotein Fc Fusion Protein Protects Guinea Pigs against Lethal Challenge.

Directory of Open Access Journals (Sweden)

Krishnamurthy Konduru

Full Text Available Ebola virus (EBOV, a member of the Filoviridae that can cause severe hemorrhagic fever in humans and nonhuman primates, poses a significant threat to the public health. Currently, there are no licensed vaccines or therapeutics to prevent and treat EBOV infection. Several vaccines based on the EBOV glycoprotein (GP are under development, including vectored, virus-like particles, and protein-based subunit vaccines. We previously demonstrated that a subunit vaccine containing the extracellular domain of the Ebola ebolavirus (EBOV GP fused to the Fc fragment of human IgG1 (EBOVgp-Fc protected mice against EBOV lethal challenge. Here, we show that the EBOVgp-Fc vaccine formulated with QS-21, alum, or polyinosinic-polycytidylic acid-poly-L-lysine carboxymethylcellulose (poly-ICLC adjuvants induced strong humoral immune responses in guinea pigs. The vaccinated animals developed anti-GP total antibody titers of approximately 105-106 and neutralizing antibody titers of approximately 103 as assessed by a BSL-2 neutralization assay based on vesicular stomatitis virus (VSV pseudotypes. The poly-ICLC formulated EBOVgp-Fc vaccine protected all the guinea pigs against EBOV lethal challenge performed under BSL-4 conditions whereas the same vaccine formulated with QS-21 or alum only induced partial protection. Vaccination with a mucin-deleted EBOVgp-Fc construct formulated with QS-21 adjuvant did not have a significant effect in anti-GP antibody levels and protection against EBOV lethal challenge compared to the full-length GP construct. The bulk of the humoral response induced by the EBOVgp-Fc vaccine was directed against epitopes outside the EBOV mucin region. Our findings indicate that different adjuvants can eliciting varying levels of protection against lethal EBOV challenge in guinea pigs vaccinated with EBOVgp-Fc, and suggest that levels of total anti-GP antibodies elicit by protein-based GP subunit vaccines do not correlate with protection. Our data

Deeper than skin deep - The effect of botulinum toxin-A on emotion processing.

Science.gov (United States)

Baumeister, J-C; Papa, G; Foroni, F

2016-08-01

The effect of facial botulinum Toxin-A (BTX) injections on the processing of emotional stimuli was investigated. The hypothesis, that BTX would interfere with processing of slightly emotional stimuli and less with very emotional or neutral stimuli, was largely confirmed. BTX-users rated slightly emotional sentences and facial expressions, but not very emotional or neutral ones, as less emotional after the treatment. Furthermore, they became slower at categorizing slightly emotional facial expressions under time pressure. Copyright © 2016 Elsevier Ltd. All rights reserved.

In vitro evaluation, biodistribution and scintigraphic imaging in mice of radiolabeled anthrax toxins

International Nuclear Information System (INIS)

Dadachova, Ekaterina; Rivera, Johanna; Revskaya, Ekaterina; Nakouzi, Antonio; Cahill, Sean M.; Blumenstein, Michael; Xiao, Hui; Rykunov, Dmitry; Casadevall, Arturo

2008-01-01

Introduction: There is a lot of interest towards creating therapies and vaccines for Bacillus anthracis, a bacterium which causes anthrax in humans and which spores can be made into potent biological weapons. Systemic injection of lethal factor (LF), edema factor (EF) and protective antigen (PA) in mice produces toxicity, and this protocol is commonly used to investigate the efficacy of specific antibodies in passive protection and vaccine studies. Availability of toxins labeled with imageable radioisotopes would allow to demonstrate their tissue distribution after intravenous injection at toxin concentration that are below pharmacologically significant to avoid masking by toxic effects. Methods: LF, EF and PA were radiolabeled with 188 Re and 99m Tc, and their performance in vitro was evaluated by macrophages and Chinese hamster ovary cells toxicity assays and by binding to macrophages. Scintigraphic imaging and biodistribution of intravenously (IV) injected 99m Tc-and 123 I-labeled toxins was performed in BALB/c mice. Results: Radiolabeled toxins preserved their biological activity. Scatchard-type analysis of the binding of radiolabeled PA to the J774.16 macrophage-like cells revealed 6.6x10 4 binding sites per cell with a dissociation constant of 6.7 nM. Comparative scintigraphic imaging of mice injected intravenously with either 99m Tc-or 123 I-labeled PA, EF and LF toxins demonstrated similar biodistribution patterns with early localization of radioactivity in the liver, spleen, intestines and excretion through kidneys. The finding of renal excretion shortly after IV injection strongly suggests that toxins are rapidly degraded which could contribute to the variability of mouse toxigenic assays. Biodistribution studies confirmed that all three toxins concentrated in the liver and the presence of high levels of radioactivity again implied rapid degradation in vivo. Conclusions: The availability of 188 Re and 99m Tc-labeled PA, LF and EF toxins allowed us to

Effect of lethal and sub-lethal concentrations of tobacco (Nicotiana ...

African Journals Online (AJOL)

Lethal and sub-lethal bioassays on Clarias gariepinus were conducted to evaluate the toxicity of tobacco (Nicotiana tobaccum) leaf dust on weight gain and haematological indices of Clarias gariepinus (mean weight 10.5±0.70g) in glass aquaria with aeration system. The concentrations used during the lethal exposure are: ...

Harvesting Venom Toxins from Assassin Bugs and Other Heteropteran Insects.

Science.gov (United States)

Walker, Andrew Allan; Rosenthal, Max; Undheim, Eivind E A; King, Glenn F

2018-04-21

Heteropteran insects such as assassin bugs (Reduviidae) and giant water bugs (Belostomatidae) descended from a common predaceous and venomous ancestor, and the majority of extant heteropterans retain this trophic strategy. Some heteropterans have transitioned to feeding on vertebrate blood (such as the kissing bugs, Triatominae; and bed bugs, Cimicidae) while others have reverted to feeding on plants (most Pentatomomorpha). However, with the exception of saliva used by kissing bugs to facilitate blood-feeding, little is known about heteropteran venoms compared to the venoms of spiders, scorpions and snakes. One obstacle to the characterization of heteropteran venom toxins is the structure and function of the venom/labial glands, which are both morphologically complex and perform multiple biological roles (defense, prey capture, and extra-oral digestion). In this article, we describe three methods we have successfully used to collect heteropteran venoms. First, we present electrostimulation as a convenient way to collect venom that is often lethal when injected into prey animals, and which obviates contamination by glandular tissue. Second, we show that gentle harassment of animals is sufficient to produce venom extrusion from the proboscis and/or venom spitting in some groups of heteropterans. Third, we describe methods to harvest venom toxins by dissection of anaesthetized animals to obtain the venom glands. This method is complementary to other methods, as it may allow harvesting of toxins from taxa in which electrostimulation and harassment are ineffective. These protocols will enable researchers to harvest toxins from heteropteran insects for structure-function characterization and possible applications in medicine and agriculture.

Purification and characterization of a phospholipase by Photobacterium damselae subsp. piscicida from cobia Rachycentron canadum.

Science.gov (United States)

Hsu, Po-Yuan; Lee, Kuo-Kau; Hu, Chih-Chuang; Liu, Ping-Chung

2014-09-01

Toxicity of the extracellular products (ECPs) and the lethal attributes of phospholipase secreted by pathogenic Photobacterium damselae subsp. piscicida from cobia Rachycentron canadum was studied. An extracellular lethal toxin in the ECPs was partially purified by using Fast Protein Liquid Chromatography system. A protein band (27 kDa) exhibited phospholipase activity on Native-PAGE (by 0.3% egg yolk agar-overlay), was excised and eluted. The pI value of the purified phospholipase was determined as 3.65 and was determined as a phospholipase C by using the Amplex™ Red phosphatidylcholine -Specific phospholipase C Assay kit. The phospholipase showed maximum activity at temperature around 4-40 °C and maximal activity at pH between 8 and 9. The enzyme was inhibited by ethylenediamine-tetraacetic acid (EDTA) and sodium dodecyl sulfate (SDS); but was activated by Ca(2+) and Mg(2+) and inactivated by Zn(2+) and Cu(2+) . Both the ECPs and phospholipase were hemolytic against erythrocytes of cobia and lethal to the fish with LD50 values of 3.25 and 0.91 µg protein g(-1) fish, respectively. In toxicity neutralization test, the rabbit antisera against the phospholipase could neutralize the toxicity of ECPs, indicating that the phospholipase is a major extracellular toxin produced by the bacterium. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Discovery of Functional Toxin/Antitoxin Systems in Bacteria by Shotgun Cloning

Energy Technology Data Exchange (ETDEWEB)

Sberro, Hila; Leavitt, Azita; Kiro, Ruth; Koh, Eugene; Peleg, Yoav; Qimron, Udi; Sorek, Rotem

2013-04-01

Toxin-antitoxin (TA) modules, composed of a toxic protein and a counteracting antitoxin, play important roles in bacterial physiology. We examined the experimental insertion of 1.5 million genes from 388 microbial genomes into an Escherichia coli host using over 8.5 million random clones. This revealed hundreds of genes (toxins) that could only be cloned when the neighboring gene (antitoxin) was present on the same clone. Clustering of these genes revealed TA families widespread in bacterial genomes, some of which deviate from the classical characteristics previously described for such modules. Introduction of these genes into E. coli validated that the toxin toxicity is mitigated by the antitoxin. Infection experiments with T7 phage showed that two of the new modules can provide resistance against phage. Moreover, our experiments revealed an 'anti-defense' protein in phage T7 that neutralizes phage resistance. Our results expose active fronts in the arms race between bacteria and phage.

Action of cholera toxin in the intestinal epithelial cells

International Nuclear Information System (INIS)

Hyun, C.S.

1982-01-01

The primary event in the action of cholera toxin on the isolated chick intestinal epithelial cell is its interaction with the cell membrane. This involves a large number (17 million per cell) of high affinity binding sites which belong to a single class. Binding of biologically active 125 I-labeled toxin is rapid, temperature-dependent, reversible, and saturable over a wide range of concentrations and includes only a small contribution from nonspecific sites. A characteristic lag phase of 10 min occurs following the complete binding of toxin before any increase in cellular cAMP levels can be detected in the isolated cells. The response (elevation of cellular cAMP) of the enterocytes to cholera toxin is linear with time for 40-50 min and causes a six- to eight-fold increase over control levels at steady stae. cAMP and agents that increase cAMP production inhibit Cl - -independent Na + influx into the isolated enterocytes whereas chlorporomazine (CPZ) which completely abolishes toxin-induced elevation of cAMP both reverses and prevents the cAMP-mediated inhibition of Na + entry. Correlation between cellular cAMP levels and the magnitude of Na + influx into the enterocytes provides evidence for a cAMP-mediated control of intestinal Na + uptake, which may represent the mechanistic basis for the antiabsorptive effect of CT and Na + during induction of intestinal secretion. The effect of cAMP on Na + but no Cl - influx in our villus cell preparation can be partially explained in terms of a cAMP-regulated Na + /H + neutral exchange system

Lethality Index 2008-2014: Less shootings, same lethality, more opacity

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Carlos Silva Forné

2017-11-01

Full Text Available This article evaluates the use of lethal force by Mexican federal security forces during shootings with presumed members of organized crime from 2008-2014. The authors use official data and press reports on deaths and wounded in shootings to construct indicators such as the number of dead civilians over the number of dead officials from the federal security forces and the number of dead civilians over the number of wounded civilians. In a context where certain factors that contribute to an excessive use of force become more common, the results of the study show a growing use of lethal force. This raises questions over the possible excessive use of lethal force as a normal or systematic practice. The study also shows a growing context of opacity in the information available to evaluate the use of lethal force and the general lack of a legal framework to regulate the use of lethal force in Mexico.

A cross-reacting material CRM197 conjugate vaccine induces diphtheria toxin neutralizing antibody response in children and adolescents infected or not with HIV.

Science.gov (United States)

Silva, Giselle P; Santos, Rafaela S; Pereira-Manfro, Wânia F; Ferreira, Bianca; Barreto, Daniella M; Frota, Ana Cristina C; Hofer, Cristina B; Milagres, Lucimar G

2017-07-05

Anti-diphtheria antibody levels decrease with aging, and frequent booster vaccinations are required to maintain herd immunity. We analyzed the diphtheria toxin neutralizing antibody (DT-Nab) response induced by a conjugate vaccine (meningococcal C polysaccharide-CRM 197 ) in HIV-vertically infected (HI) children and adolescents and healthy controls (HC) with matched age. We report the association of DT-Nab with the bactericidal antibodies to serogroup C meningococcus (MenC). Before vaccination, 21 HI patients (50%) had no protection against diphtheria (≤0.01IU/ml of antibody) and only 8 (19%) showed complete protection (≥0.1IU/ml). About half of the HC (56%) had complete protection before immunization and 6 subjects (12%) had no protection against diphtheria. After one and two vaccine injections, 96% of HC and 64% of HI vaccinees, respectively, showed full protection against diphtheria. These data indicate that CRM 197 was able to induce primary and/or booster response in both groups of individuals. Copyright © 2017 Elsevier Ltd. All rights reserved.

Stool C difficile toxin

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... toxin; Colitis - toxin; Pseudomembranous - toxin; Necrotizing colitis - toxin; C difficile - toxin ... be analyzed. There are several ways to detect C difficile toxin in the stool sample. Enzyme immunoassay ( ...

Structure, Biology, and Therapeutic Application of Toxin-Antitoxin Systems in Pathogenic Bacteria.

Science.gov (United States)

Lee, Ki-Young; Lee, Bong-Jin

2016-10-22

Bacterial toxin-antitoxin (TA) systems have received increasing attention for their diverse identities, structures, and functional implications in cell cycle arrest and survival against environmental stresses such as nutrient deficiency, antibiotic treatments, and immune system attacks. In this review, we describe the biological functions and the auto-regulatory mechanisms of six different types of TA systems, among which the type II TA system has been most extensively studied. The functions of type II toxins include mRNA/tRNA cleavage, gyrase/ribosome poison, and protein phosphorylation, which can be neutralized by their cognate antitoxins. We mainly explore the similar but divergent structures of type II TA proteins from 12 important pathogenic bacteria, including various aspects of protein-protein interactions. Accumulating knowledge about the structure-function correlation of TA systems from pathogenic bacteria has facilitated a novel strategy to develop antibiotic drugs that target specific pathogens. These molecules could increase the intrinsic activity of the toxin by artificially interfering with the intermolecular network of the TA systems.

Detection and effects of harmful algal toxins in Scottish harbour seals and potential links to population decline.

Science.gov (United States)

Jensen, Silje-Kristin; Lacaze, Jean-Pierre; Hermann, Guillaume; Kershaw, Joanna; Brownlow, Andrew; Turner, Andrew; Hall, Ailsa

2015-04-01

Over the past 15 years or so, several Scottish harbour seal (Phoca vitulina) populations have declined in abundance and several factors have been considered as possible causes, including toxins from harmful algae. Here we explore whether a link could be established between two groups of toxins, domoic acid (DA) and saxitoxins (STXs), and the decline in the harbour seal populations in Scotland. We document the first evidence that harbour seals are exposed to both DA and STXs from consuming contaminated fish. Both groups of toxins were found in urine and faeces sampled from live captured (n = 162) and stranded animals (n = 23) and in faecal samples collected from seal haul-out sites (n = 214) between 2008 and 2013. The proportion of positive samples and the toxins levels measured in the excreta were significantly higher in areas where harbour seal abundance is in decline. There is also evidence that DA has immunomodulatory effects in harbour seals, including lymphocytopenia and monocytosis. Scottish harbour seals are exposed to DA and STXs through contaminated prey at potentially lethal levels and with this evidence we suggest that exposure to these toxins are likely to be important factors driving the harbour seal decline in some regions of Scotland. Copyright © 2015 Elsevier Ltd. All rights reserved.

Cellular vacuoles induced by Mycoplasma pneumoniae CARDS toxin originate from Rab9-associated compartments.

Directory of Open Access Journals (Sweden)

Coreen Johnson

Full Text Available Recently, we identified an ADP-ribosylating and vacuolating cytotoxin in Mycoplasma pneumoniae designated Community Acquired Respiratory Distress Syndrome (CARDS toxin. In this study we show that vacuoles induced by recombinant CARDS (rCARDS toxin are acidic and derive from the endocytic pathway as determined by the uptake of neutral red and the fluid-phase marker, Lucifer yellow, respectively. Also, we demonstrate that the formation of rCARDS toxin-associated cytoplasmic vacuoles is inhibited by the vacuolar ATPase inhibitor, bafilomycin A1, and the ionophore, monensin. To examine the ontogeny of these vacuoles, we analyzed the distribution of endosomal and lysosomal membrane markers during vacuole formation and observed the enrichment of the late endosomal GTPase, Rab9, around rCARDS toxin-induced vacuoles. Immunogold-labeled Rab9 and overexpression of green fluorescent-tagged Rab9 further confirmed vacuolar association. The late endosomal- and lysosomal-associated membrane proteins, LAMP1 and LAMP2, also localized to the vacuolar membranes, while the late endosomal protein, Rab7, and early endosomal markers, Rab5 and EEA1, were excluded. HeLa cells expressing dominant-negative (DN Rab9 exhibited markedly reduced vacuole formation in the presence of rCARDS toxin, in contrast to cells expressing DN-Rab7, highlighting the importance of Rab9 function in rCARDS toxin-induced vacuolation. Our findings reveal the unique Rab9-association with rCARDS toxin-induced vacuoles and its possible relationship to the characteristic histopathology that accompanies M. pneumoniae infection.

Immunization with cholera toxin B subunit induces high-level protection in the suckling mouse model of cholera.

Directory of Open Access Journals (Sweden)

Gregory A Price

Full Text Available Cholera toxin (CT is the primary virulence factor responsible for severe cholera. Vibrio cholerae strains unable to produce CT show severe attenuation of virulence in animals and humans. The pentameric B subunit of CT (CTB contains the immunodominant epitopes recognized by antibodies that neutralize CT. Although CTB is a potent immunogen and a promising protective vaccine antigen in animal models, immunization of humans with detoxified CT failed to protect against cholera. We recently demonstrated however that pups reared from mice immunized intraperitoneally (IP with 3 doses of recombinant CTB were well protected against a highly lethal challenge dose of V. cholerae N16961. The present study investigated how the route and number of immunizations with CTB could influence protective efficacy in the suckling mouse model of cholera. To this end female mice were immunized with CTB intranasally (IN, IP, and subcutaneously (SC. Serum and fecal extracts were analyzed for anti-CTB antibodies by quantitative ELISA, and pups born to immunized mothers were challenged orogastrically with a lethal dose of V. cholerae. Pups from all immunized groups were highly protected from death by 48 hours (64-100% survival. Cox regression showed that percent body weight loss at 24 hours predicted death by 48 hours, but we were unable to validate a specific amount of weight loss as a surrogate marker for protection. Although CTB was highly protective in all regimens, three parenteral immunizations showed trends toward higher survival and less weight loss at 24 hours post infection. These results demonstrate that immunization with CTB by any of several routes and dosing regimens can provide protection against live V. cholerae challenge in the suckling mouse model of cholera. Our data extend the results of previous studies and provide additional support for the inclusion of CTB in the development of a subunit vaccine against V. cholerae.

Mechanistic insights into the neutralization of cytotoxic abrin by the monoclonal antibody D6F10.

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Shradha Bagaria

Full Text Available Abrin, an A/B toxin obtained from the Abrus precatorius plant is extremely toxic and a potential bio-warfare agent. Till date there is no antidote or vaccine available against this toxin. The only known neutralizing monoclonal antibody against abrin, namely D6F10, has been shown to rescue the toxicity of abrin in cells as well as in mice. The present study focuses on mapping the epitopic region to understand the mechanism of neutralization of abrin by the antibody D6F10. Truncation and mutational analysis of abrin A chain revealed that the amino acids 74-123 of abrin A chain contain the core epitope and the residues Thr112, Gly114 and Arg118 are crucial for binding of the antibody. In silico analysis of the position of the mapped epitope indicated that it is present close to the active site cleft of abrin A chain. Thus, binding of the antibody near the active site blocks the enzymatic activity of abrin A chain, thereby rescuing inhibition of protein synthesis by the toxin in vitro. At 1∶10 molar concentration of abrin:antibody, the antibody D6F10 rescued cells from abrin-mediated inhibition of protein synthesis but did not prevent cell attachment of abrin. Further, internalization of the antibody bound to abrin was observed in cells by confocal microscopy. This is a novel finding which suggests that the antibody might function intracellularly and possibly explains the rescue of abrin's toxicity by the antibody in whole cells and animals. To our knowledge, this study is the first report on a neutralizing epitope for abrin and provides mechanistic insights into the poorly understood mode of action of anti-A chain antibodies against several toxins including ricin.

Enterotoxigenic Escherichia coli Adhesin-Toxoid Multiepitope Fusion Antigen CFA/I/II/IV-3xSTaN12S-mnLTG192G/L211A-Derived Antibodies Inhibit Adherence of Seven Adhesins, Neutralize Enterotoxicity of LT and STa Toxins, and Protect Piglets against Diarrhea.

Science.gov (United States)

Nandre, Rahul; Ruan, Xiaosai; Lu, Ti; Duan, Qiangde; Sack, David; Zhang, Weiping

2018-03-01

Enterotoxigenic Escherichia coli (ETEC) strains are a leading cause of children's diarrhea and travelers' diarrhea. Vaccines inducing antibodies to broadly inhibit bacterial adherence and to neutralize toxin enterotoxicity are expected to be effective against ETEC-associated diarrhea. 6×His-tagged adhesin-toxoid fusion proteins were shown to induce neutralizing antibodies to several adhesins and LT and STa toxins (X. Ruan, D. A. Sack, W. Zhang, PLoS One 10:e0121623, 2015, https://doi.org/10.1371/journal.pone.0121623). However, antibodies derived from His-tagged CFA/I/II/IV-2xSTa A14Q -dmLT or CFA/I/II/IV-2xSTa N12S -dmLT protein were less effective in neutralizing STa enterotoxicity and were not evaluated in vivo for efficacy against ETEC diarrhea. Additionally, His-tagged proteins are considered less desirable for human vaccines. In this study, we produced a tagless adhesin-toxoid MEFA (multiepitope fusion antigen) protein, enhanced anti-STa immunogenicity by including a third copy of STa toxoid STa N12S , and examined antigen immunogenicity in a murine model. Moreover, we immunized pregnant pigs with the tagless adhesin-toxoid MEFA protein and evaluated passive antibody protection against STa + or LT + ETEC infection in a pig challenge model. Results showed that tagless adhesin-toxoid MEFA CFA/I/II/IV-3xSTa N12S -mnLT R192G/L211A induced broad antiadhesin and antitoxin antibody responses in the intraperitoneally immunized mice and the intramuscularly immunized pigs. Mouse and pig serum antibodies significantly inhibited adherence of seven colonization factor antigen (CFA) adhesins (CFA/I and CS1 to CS6) and effectively neutralized both toxins. More importantly, suckling piglets born to the immunized mothers acquired antibodies and were protected against STa + ETEC and LT + ETEC diarrhea. These results indicated that tagless CFA/I/II/IV-3xSTa N12S -mnLT R192G/L211A induced broadly protective antiadhesin and antitoxin antibodies and demonstrate that this adhesin

Ligand-induced expansion of the S1' site in the anthrax toxin lethal factor

Energy Technology Data Exchange (ETDEWEB)

Maize, Kimberly M.; Kurbanov, Elbek K.; Johnson, Rodney L.; Amin, Elizabeth Ambrose; Finzel, Barry C. (UMM)

2016-07-05

The Bacillus anthracis lethal factor (LF) is one component of a tripartite exotoxin partly responsible for persistent anthrax cytotoxicity after initial bacterial infection. Inhibitors of the zinc metalloproteinase have been investigated as potential therapeutic agents, but LF is a challenging target because inhibitors lack sufficient selectivity or possess poor pharmaceutical properties. These structural studies reveal an alternate conformation of the enzyme, induced upon binding of specific inhibitors, that opens a previously unobserved deep pocket termed S1'* which might afford new opportunities to design selective inhibitors that target this subsite.

Histopathological effects of lethal and sub-lethal concentrations of ...

African Journals Online (AJOL)

The histopathological effects of lethal and sub-lethal concentrations of glyphosate on African catfish Clarias gariepinus were investigated. C. gariepinus juveniles were assessed in a static renewal bioassay for 96 hours (acute toxicity) and 28 days (chronic toxicity) using varying concentrations (0.0 mg/l 20.0 mg/l, 30.0 mg/l, ...

Ciprofloxacin causes persister formation by inducing the TisB toxin in Escherichia coli.

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Tobias Dörr

2010-02-01

Full Text Available Bacteria induce stress responses that protect the cell from lethal factors such as DNA-damaging agents. Bacterial populations also form persisters, dormant cells that are highly tolerant to antibiotics and play an important role in recalcitrance of biofilm infections. Stress response and dormancy appear to represent alternative strategies of cell survival. The mechanism of persister formation is unknown, but isolated persisters show increased levels of toxin/antitoxin (TA transcripts. We have found previously that one or more components of the SOS response induce persister formation after exposure to a DNA-damaging antibiotic. The SOS response induces several TA genes in Escherichia coli. Here, we show that a knockout of a particular SOS-TA locus, tisAB/istR, had a sharply decreased level of persisters tolerant to ciprofloxacin, an antibiotic that causes DNA damage. Step-wise administration of ciprofloxacin induced persister formation in a tisAB-dependent manner, and cells producing TisB toxin were tolerant to multiple antibiotics. TisB is a membrane peptide that was shown to decrease proton motive force and ATP levels, consistent with its role in forming dormant cells. These results suggest that a DNA damage-induced toxin controls production of multidrug tolerant cells and thus provide a model of persister formation.

Inhibition of cholera toxin and other AB toxins by polyphenolic compounds

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All AB-type protein toxins have intracellular targets despite an initial extracellular location. These toxins use different methods to reach the cytosol and have different effects on the target cell. Broad-spectrum inhibitors against AB toxins are therefore hard to develop because the toxins use dif...

Anti-lipopolysaccharide toxin therapy for whole body X-irradiation overdose

Energy Technology Data Exchange (ETDEWEB)

Gaffin, S.L.; Wells, M.; Jordan, J.P.

1985-09-01

Death in humans from ionising radiation overexposure in the 3-8 Gy (300-800 rad) range is in part due to the toxaemia caused by the entry of gram-negative bacteria and/or their lipopolysaccharide toxin (LPS) into the blood circulation through the walls of partially denuded gut. Anti-LPS hyperimmune equine plasma was evaluated for its ability to lower irradiation-induced lethality. Mice were irradiated with 6.3 Gy (630 rad) and six days later received equine Anti-LPS hyperimmune plasma, control plasma or saline. Mortalities in the three groups were 58%, 92% and 79% (p < 0.01) respectively. Thus Anti-LPS may prove useful as an adjunct to conventional therapy in treating radiation sickness.

Anti-lipopolysaccharide toxin therapy for whole body X-irradiation overdose

International Nuclear Information System (INIS)

Gaffin, S.L.; Wells, M.; Jordan, J.P.

1985-01-01

Death in humans from ionising radiation overexposure in the 3-8 Gy (300-800 rad) range is in part due to the toxaemia caused by the entry of gram-negative bacteria and/or their lipopolysaccharide toxin (LPS) into the blood circulation through the walls of partially denuded gut. Anti-LPS hyperimmune equine plasma was evaluated for its ability to lower irradiation-induced lethality. Mice were irradiated with 6.3 Gy (630 rad) and six days later received equine Anti-LPS hyperimmune plasma, control plasma or saline. Mortalities in the three groups were 58%, 92% and 79% (p<0.01) respectively. Thus Anti-LPS may prove useful as an adjunct to conventional therapy in treating radiation sickness. (author)

Crystallization and preliminary X-ray analysis of the vWA domain of human anthrax toxin receptor 1

International Nuclear Information System (INIS)

Cai, Chenguang; Zhao, Ying; Tong, Xiaohang; Fu, Sheng; Li, Yuanyuan; Wu, Yang; Li, Xumei; Lou, Zhiyong

2010-01-01

The vWA domain of human anthrax toxin receptor 1 was overexpressed in E. coli, purified and crystallized. Diffraction data were collected to 1.8 Å resolution. The Gram-positive spore-forming bacterium Bacillus anthracis causes anthrax by secreting anthrax toxin, which consists of protective antigen (PA), lethal factor and oedema factor. Binding of PA to receptors triggers the multi-step process of anthrax toxin entry into target cells. Two distinct cellular receptors, ANTXR1 (also known as tumour endothelial marker 8; TEM8) and ANTXR2 (also known as capillary morphogenesis protein 2; CMG2), for anthrax toxin have been identified. Although the crystal structure of the extracellular von Willebrand factor A (vWA) domain of CMG2 has been reported, the difference between the vWA domains of TEM8 and CMG2 remains unclear because there are no structural data for the TEM8 vWA domain. In this report, the TEM8 vWA domain was expressed, purified and crystallized. X-ray diffraction data were collected to 1.8 Å resolution from a single crystal, which belonged to space group P1 with unit-cell parameters a = 65.9, b = 66.1, c = 74.4 Å, α = 63.7, β = 88.2, γ = 59.9°

Tolerization with BLP down-regulates HMGB1 a critical mediator of sepsis-related lethality.

LENUS (Irish Health Repository)

Coffey, J Calvin

2012-02-03

Tolerization with bacterial lipoprotein (BLP) affords a significant survival benefit in sepsis. Given that high mobility group box protein-1 (HMGB1) is a recognized mediator of sepsis-related lethality, we determined if tolerization with BLP leads to alterations in HMGB1. In vitro, BLP tolerization led to a reduction in HMGB1 gene transcription. This was mirrored at the protein level, as HMGB1 protein expression and release were reduced significantly in BLP-tolerized human THP-1 monocytic cells. BLP tolerance in vivo led to a highly significant, long-term survival benefit following challenge with lethal dose BLP in C57BL\\/6 mice. This was associated with an attenuation of HMGB1 release into the circulation, as evidenced by negligible serum HMGB1 levels in BLP-tolerized mice. Moreover, HMGB1 levels in peritoneal macrophages from BLP-tolerized mice were reduced significantly. Hence, tolerization with BLP leads to a down-regulation of HMGB1 protein synthesis and release. The improved survival associated with BLP tolerance could thus be explained by a reduction in HMGB1, were the latter associated with lethality in BLP-related sepsis. In testing this hypothesis, it was noted that neutralization of HMGB1, using anti-HMGB1 antibodies, abrogated BLP-associated lethality almost completely. To conclude, tolerization with BLP leads to a down-regulation of HMGB1, thus offering a novel means of targeting the latter. HMGB1 is also a mediator of lethality in BLP-related sepsis.

Anthrax lethal factor as an immune target in humans and transgenic mice and the impact of HLA polymorphism on CD4+ T cell immunity.

Science.gov (United States)

Ascough, Stephanie; Ingram, Rebecca J; Chu, Karen K; Reynolds, Catherine J; Musson, Julie A; Doganay, Mehmet; Metan, Gökhan; Ozkul, Yusuf; Baillie, Les; Sriskandan, Shiranee; Moore, Stephen J; Gallagher, Theresa B; Dyson, Hugh; Williamson, E Diane; Robinson, John H; Maillere, Bernard; Boyton, Rosemary J; Altmann, Daniel M

2014-05-01

Bacillus anthracis produces a binary toxin composed of protective antigen (PA) and one of two subunits, lethal factor (LF) or edema factor (EF). Most studies have concentrated on induction of toxin-specific antibodies as the correlate of protective immunity, in contrast to which understanding of cellular immunity to these toxins and its impact on infection is limited. We characterized CD4+ T cell immunity to LF in a panel of humanized HLA-DR and DQ transgenic mice and in naturally exposed patients. As the variation in antigen presentation governed by HLA polymorphism has a major impact on protective immunity to specific epitopes, we examined relative binding affinities of LF peptides to purified HLA class II molecules, identifying those regions likely to be of broad applicability to human immune studies through their ability to bind multiple alleles. Transgenics differing only in their expression of human HLA class II alleles showed a marked hierarchy of immunity to LF. Immunogenicity in HLA transgenics was primarily restricted to epitopes from domains II and IV of LF and promiscuous, dominant epitopes, common to all HLA types, were identified in domain II. The relevance of this model was further demonstrated by the fact that a number of the immunodominant epitopes identified in mice were recognized by T cells from humans previously infected with cutaneous anthrax and from vaccinated individuals. The ability of the identified epitopes to confer protective immunity was demonstrated by lethal anthrax challenge of HLA transgenic mice immunized with a peptide subunit vaccine comprising the immunodominant epitopes that we identified.

Some properties and cDNA cloning of proteinaceous toxins from two species of lionfish (Pterois antennata and Pterois volitans).

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Kiriake, Aya; Shiomi, Kazuo

2011-11-01

Lionfish, members of the genera Pterois, Parapterois and Dendrochirus, are well known to be venomous, having venomous glandular tissues in dorsal, pelvic and anal spines. The lionfish toxins have been shown to cross-react with the stonefish toxins by neutralization tests using the commercial stonefish antivenom, although their chemical properties including structures have been little characterized. In this study, an antiserum against neoverrucotoxin, the stonefish Synanceia verrucosa toxin, was first raised in a guinea pig and used in immunoblotting and inhibition immunoblotting to confirm that two species of Pterois lionfish (P. antennata and P. volitans) contain a 75kDa protein (corresponding to the toxin subunit) cross-reacting with neoverrucotoxin. Then, the amino acid sequences of the P. antennata and P. volitans toxins were successfully determined by cDNA cloning using primers designed from the highly conserved sequences of the stonefish toxins. Notably, either α-subunits (699 amino acid residues) or β-subunits (698 amino acid residues) of the P. antennata and P. volitans toxins share as high as 99% sequence identity with each other. Furthermore, both α- and β-subunits of the lionfish toxins exhibit high sequence identity (70-80% identity) with each other and also with the β-subunits of the stonefish toxins. As reported for the stonefish toxins, the lionfish toxins also contain a B30.2/SPRY domain (comprising nearly 200 amino acid residues) in the C-terminal region of each subunit. Copyright © 2011 Elsevier Ltd. All rights reserved.

Binding of ATP by pertussis toxin and isolated toxin subunits

International Nuclear Information System (INIS)

Hausman, S.Z.; Manclark, C.R.; Burns, D.L.

1990-01-01

The binding of ATP to pertussis toxin and its components, the A subunit and B oligomer, was investigated. Whereas, radiolabeled ATP bound to the B oligomer and pertussis toxin, no binding to the A subunit was observed. The binding of [ 3 H]ATP to pertussis toxin and the B oligomer was inhibited by nucleotides. The relative effectiveness of the nucleotides was shown to be ATP > GTP > CTP > TTP for pertussis toxin and ATP > GTP > TTP > CTP for the B oligomer. Phosphate ions inhibited the binding of [ 3 H]ATP to pertussis toxin in a competitive manner; however, the presence of phosphate ions was essential for binding of ATP to the B oligomer. The toxin substrate, NAD, did not affect the binding of [ 3 H]ATP to pertussis toxin, although the glycoprotein fetuin significantly decreased binding. These results suggest that the binding site for ATP is located on the B oligomer and is distinct from the enzymatically active site but may be located near the eukaryotic receptor binding site

Binding of ATP by pertussis toxin and isolated toxin subunits

Energy Technology Data Exchange (ETDEWEB)

Hausman, S.Z.; Manclark, C.R.; Burns, D.L. (Center for Biologics Evaluation and Research, Bethesda, MD (USA))

1990-07-03

The binding of ATP to pertussis toxin and its components, the A subunit and B oligomer, was investigated. Whereas, radiolabeled ATP bound to the B oligomer and pertussis toxin, no binding to the A subunit was observed. The binding of ({sup 3}H)ATP to pertussis toxin and the B oligomer was inhibited by nucleotides. The relative effectiveness of the nucleotides was shown to be ATP > GTP > CTP > TTP for pertussis toxin and ATP > GTP > TTP > CTP for the B oligomer. Phosphate ions inhibited the binding of ({sup 3}H)ATP to pertussis toxin in a competitive manner; however, the presence of phosphate ions was essential for binding of ATP to the B oligomer. The toxin substrate, NAD, did not affect the binding of ({sup 3}H)ATP to pertussis toxin, although the glycoprotein fetuin significantly decreased binding. These results suggest that the binding site for ATP is located on the B oligomer and is distinct from the enzymatically active site but may be located near the eukaryotic receptor binding site.

Antitoxin activity of aqueous extract of Cyclea peltata root against Naja naja venom.

Science.gov (United States)

Sivaraman, Thulasi; Sreedevi, N S; Meenatchisundaram, S; Vadivelan, R

2017-01-01

Snakebites are a significant and severe global health problem. Till date, anti-snake venom serum is the only beneficial remedy existing on treating the snakebite victims. As antivenom was reported to induce early or late adverse reactions to human beings, snake venom neutralizing potential for Cyclea peltata root extract was tested for the present research by ex vivo and in vivo approaches on Naja naja toxin. Ex vivo evaluation of venom toxicity and neutralization assays was carried out. The root extracts from C. peltata were used to evaluate the Ex vivo neutralization tests such as acetylcholinesterase, protease, direct hemolysis assay, phospholipase activity, and procoagulant activity. Gas chromatography-mass spectrometry (GC-MS) analysis from root extracts of C. peltata was done to investigate the bioactive compounds. The in vivo calculation of venom toxicity (LD 50 ) of N. naja venom remained to be 0.301 μg. C. peltata root extracts were efficiently deactivated the venom lethality, and effective dose (ED 50 ) remained to be 7.24 mg/3LD 50 of N. naja venom. C. peltata root extract was found effective in counteracting all the lethal effects of venom. GC-MS analysis of the plant extract revealed the presence of antivenom compounds such as tetradecanoic and octadecadienoic acid which have neutralizing properties on N. naja venom. The result from the ex vivo and in vivo analysis indicates that C. peltata plant root extract possesses significant compounds such as tetradecanoic acid hexadecanoic acid, heptadecanoic acid, and octadecadienoic acid which can counteract the toxins present in N. naja .

Tumor endothelium marker-8 based decoys exhibit superiority over capillary morphogenesis protein-2 based decoys as anthrax toxin inhibitors.

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Chenguang Cai

Full Text Available Anthrax toxin is the major virulence factor produced by Bacillus anthracis. The toxin consists of three protein subunits: protective antigen (PA, lethal factor, and edema factor. Inhibition of PA binding to its receptors, tumor endothelium marker-8 (TEM8 and capillary morphogenesis protein-2 (CMG2 can effectively block anthrax intoxication, which is particularly valuable when the toxin has already been overproduced at the late stage of anthrax infection, thus rendering antibiotics ineffectual. Receptor-like agonists, such as the mammalian cell-expressed von Willebrand factor type A (vWA domain of CMG2 (sCMG2, have demonstrated potency against the anthrax toxin. However, the soluble vWA domain of TEM8 (sTEM8 was ruled out as an anthrax toxin inhibitor candidate due to its inferior affinity to PA. In the present study, we report that L56A, a PA-binding-affinity-elevated mutant of sTEM8, could inhibit anthrax intoxication as effectively as sCMG2 in Fisher 344 rats. Additionally, pharmacokinetics showed that L56A and sTEM8 exhibit advantages over sCMG2 with better lung-targeting and longer plasma retention time, which may contribute to their enhanced protective ability in vivo. Our results suggest that receptor decoys based on TEM8 are promising anthrax toxin inhibitors and, together with the pharmacokinetic studies in this report, may contribute to the development of novel anthrax drugs.

Toxin-antitoxin systems and regulatory mechanisms in Mycobacterium tuberculosis.

Science.gov (United States)

Slayden, Richard A; Dawson, Clinton C; Cummings, Jason E

2018-06-01

There has been a significant reduction in annual tuberculosis incidence since the World Health Organization declared tuberculosis a global health threat. However, treatment of M. tuberculosis infections requires lengthy multidrug therapeutic regimens to achieve a durable cure. The development of new drugs that are active against resistant strains and phenotypically diverse organisms continues to present the greatest challenge in the future. Numerous phylogenomic analyses have revealed that the Mtb genome encodes a significantly expanded repertoire of toxin-antitoxin (TA) loci that makes up the Mtb TA system. A TA loci is a two-gene operon encoding a 'toxin' protein that inhibits bacterial growth and an interacting 'antitoxin' partner that neutralizes the inhibitory activity of the toxin. The presence of multiple chromosomally encoded TA loci in Mtb raises important questions in regard to expansion, regulation and function. Thus, the functional roles of TA loci in Mtb pathogenesis have received considerable attention over the last decade. The cumulative results indicate that they are involved in regulating adaptive responses to stresses associated with the host environment and drug treatment. Here we review the TA families encoded in Mtb, discuss the duplication of TA loci in Mtb, regulatory mechanism of TA loci, and phenotypic heterogeneity and pathogenesis.

Alpha-beta T cells provide protection against lethal encephalitis in the murine model of VEEV infection

International Nuclear Information System (INIS)

Paessler, Slobodan; Yun, Nadezhda E.; Judy, Barbara M.; Dziuba, Natallia; Zacks, Michele A.; Grund, Anna H.; Frolov, Ilya; Campbell, Gerald A.; Weaver, Scott C.; Estes, D. Mark

2007-01-01

We evaluated the safety and immunogenicity of a chimeric alphavirus vaccine candidate in mice with selective immunodeficiencies. This vaccine candidate was highly attenuated in mice with deficiencies in the B and T cell compartments, as well as in mice with deficient gamma-interferon responsiveness. However, the level of protection varied among the strains tested. Wild type mice were protected against lethal VEEV challenge. In contrast, alpha/beta (αβ) TCR-deficient mice developed lethal encephalitis following VEEV challenge, while mice deficient in gamma/delta (γδ) T cells were protected. Surprisingly, the vaccine potency was diminished by 50% in animals lacking interferon-gamma receptor alpha chain (R1)-chain and a minority of vaccinated immunoglobulin heavy chain-deficient (μMT) mice survived challenge, which suggests that neutralizing antibody may not be absolutely required for protection. Prolonged replication of encephalitic VEEV in the brain of pre-immunized mice is not lethal and adoptive transfer experiments indicate that CD3 + T cells are required for protection

Unveiling the nature of black mamba (Dendroaspis polylepis) venom through venomics and antivenom immunoprofiling: Identification of key toxin targets for antivenom development

DEFF Research Database (Denmark)

Laustsen, Andreas Hougaard; Lomonte, Bruno; Lohse, Brian

2015-01-01

The venom proteome of the black mamba, Dendroaspis polylepis, from Eastern Africa, was, for the first time, characterized. Forty- different proteins and one nucleoside were identified or assigned to protein families. The most abundant proteins were Kunitz-type proteinase inhibitors, which include...... the unique mamba venom components ‘dendrotoxins’, and α-neurotoxins and other representatives of the three-finger toxin family. In addition, the venom contains lower percentages of proteins from other families, including metalloproteinase, hyaluronidase, prokineticin, nerve growth factor, vascular...... to toxicity by influencing the toxin biodistribution. ELISA immunoprofiling and preclinical assessment of neutralization showed that polyspecific antivenoms manufactured in South Africa and India were effective in the neutralization of D. polylepis venom, albeit showing different potencies. Antivenoms had...

SINGLE CHAIN VARIABLE FRAGMENTS OF ANTIBODIES AGAINST DIPHTHERIA TOXIN B-SUBUNIT ISOLATED FROM PHAGE DISPLAY HUMAN ANTIBODY LIBRARY

Directory of Open Access Journals (Sweden)

Oliinyk O. S.

2014-02-01

Full Text Available Diphtheria toxin is an exoantigen of Corynebacterium diphtheriae that inhibits protein synthesis and kills sensitive cells. The aim of this study was to obtain human recombinant single-chain variable fragment (scFv antibodies against receptor-binding B subunit of diphtheria toxin. 12 specific clones were selected after three rounds of a phage display naїve (unimmunized human antibody library against recombinant B-subunit. scFv DNA inserts from these 12 clones were digested with MvaI, and 6 unique restriction patterns were found. Single-chain antibodies were expressed in Escherichia coli XL1-blue. The recombinant proteins were characterized by immunoblotting of bacterial extracts and detection with an anti-E-tag antibody. The toxin B-subunit-binding function of the single-chain antibody was shown by ELISA. The affinity constants for different clones were found to be from 106 to 108 М–1. Due to the fact, that these antibody fragments recognized epitopes in the receptor-binding Bsubunit of diphtheria toxin, further studies are interesting to evaluate their toxin neutralization properties and potential for therapeutic applications. Obtained scFv-antibodies can also be used for detection and investigation of biological properties of diphtheria toxin.

Isolation of Anti-Ricin Protective Antibodies Exhibiting High Affinity from Immunized Non-Human Primates

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Tal Noy-Porat

2016-03-01

Full Text Available Ricin, derived from the castor bean plant Ricinus communis, is one of the most potent and lethal toxins known, against which there is no available antidote. To date, the use of neutralizing antibodies is the most promising post-exposure treatment for ricin intoxication. The aim of this study was to isolate high affinity anti-ricin antibodies that possess potent toxin-neutralization capabilities. Two non-human primates were immunized with either a ricin-holotoxin- or subunit-based vaccine, to ensure the elicitation of diverse high affinity antibodies. By using a comprehensive set of primers, immune scFv phage-displayed libraries were constructed and panned. A panel of 10 antibodies (five directed against the A subunit of ricin and five against the B subunit was isolated and reformatted into a full-length chimeric IgG. All of these antibodies were found to neutralize ricin in vitro, and several conferred full protection to ricin-intoxicated mice when given six hours after exposure. Six antibodies were found to possess exceptionally high affinity toward the toxin, with KD values below pM (koff < 1 × 10−7 s−1 that were well correlated with their ability to neutralize ricin. These antibodies, alone or in combination, could be used for the development of a highly-effective therapeutic preparation for post-exposure treatment of ricin intoxication.

Linking ciguatera poisoning to spatial ecology of fish: a novel approach to examining the distribution of biotoxin levels in the great barracuda by combining non-lethal blood sampling and biotelemetry.

Science.gov (United States)

O'Toole, Amanda C; Dechraoui Bottein, Marie-Yasmine; Danylchuk, Andy J; Ramsdell, John S; Cooke, Steven J

2012-06-15

Ciguatera in humans is typically caused by the consumption of reef fish that have accumulated Ciguatoxins (CTXs) in their flesh. Over a six month period, we captured 38 wild adult great barracuda (Sphyraena barracuda), a species commonly associated with ciguatera in The Bahamas. We sampled three tissues (i.e., muscle, liver, and blood) and analysed them for the presence of ciguatoxins using a functional in vitro N2A bioassay. Detectable concentrations of ciguatoxins found in the three tissue types ranged from 2.51 to 211.74pg C-CTX-1 equivalents/g. Blood and liver toxin concentrations were positively correlated (ρ=0.86, P=0.003), indicating that, for the first time, blood sampling provides a non-lethal method of detecting ciguatoxin in wild fish. Non-lethal blood sampling also presents opportunities to couple this approach with biotelemetry and biologging techniques that enable the study of fish distribution and movement. To demonstrate the potential for linking ciguatoxin occurrence with barracuda spatial ecology, we also present a proof-of-concept case study where blood samples were obtained from 20 fish before releasing them with acoustic transmitters and tracking them in the coastal waters using a fixed acoustic telemetry array covering 44km(2). Fish that tested positive for CTX may have smaller home ranges than non-toxic fish (median distance travelled, U=2.21, P=0.03). Results presented from this study may help identify high risk areas and source-sink dynamics of toxins, potentially reducing the incidence and human health risk of ciguatera fish poisoning. Moreover, development of the non-lethal sampling approach and measurement of ciguatera from blood provide future opportunities to understand the mechanistic relationship between toxins and the spatial ecology of a broad range of marine fish species. Copyright © 2012 Elsevier B.V. All rights reserved.

Immunogenicity test of tetanus component in adsorbed vaccines by toxin binding inhibition test

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Denise Cristina Souza Matos

2002-09-01

Full Text Available Samples from 20 lots of diphtheria-tetanus (adult use dT vaccine and from 20 lots of diphtheria-tetanus-pertussis (DTP vaccine were used to standardize and validate the in vitro toxin binding inhibition (ToBI test for the immunogenicity test of the tetanus component. The levels of tetanus antitoxin obtained by ToBI test were compared to those obtained using the toxin neutralization (TN test in mice routinely employed to perform the quality control of the tetanus component in adsorbed vaccines. The results ranged from 1.8 to 3.5 IU/ml for dT and 2 to 4 IU/ml for DTP by ToBI test and 1.4 to 3 IU/ml for dT and 1.8 to 3.5 IU/ml for DTP by TN in mice. These results were significantly correlated. From this study, it is concluded that the ToBI test is an alternative to the in vivo neutralization procedure in the immunogenicity test of the tetanus component in adsorbed vaccines. A substantial refinement and a reduction in use of animals can be achieved.

Failure of botulinum toxin injection for neurogenic detrusor overactivity: Switch of toxin versus second injection of the same toxin.

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Peyronnet, Benoit; Castel-Lacanal, Evelyne; Manunta, Andréa; Roumiguié, Mathieu; Marque, Philippe; Rischmann, Pascal; Gamé, Xavier

2015-12-01

To evaluate the efficacy of a second injection of the same toxin versus switching to a different botulinum toxin A after failure of a first detrusor injection in patients with neurogenic detrusor overactivity. The charts of all patients who underwent detrusor injections of botulinum toxin A (either abobotulinumtoxinA or onabotulinumtoxinA) for the management of neurogenic detrusor overactivity at a single institution were retrospectively reviewed. Patients in whom a first detrusor injection had failed were included in the present study. They were managed by a second injection of the same toxin at the same dosage or by a new detrusor injection using a different botulinum toxin A. Success was defined as a resolution of urgency, urinary incontinence and detrusor overactivity in a patient self-catheterizing seven times or less per 24 h. A total of 58 patients were included for analysis. A toxin switch was carried out in 29 patients, whereas the other 29 patients received a reinjection of the same toxin at the same dose. The success rate was higher in patients who received a toxin switch (51.7% vs. 24.1%, P = 0.03). Patients treated with a switch from abobotulinumtoxinA to onabotulinumtoxinA and those treated with a switch from onabotulinumtoxinA to abobotulinumtoxinA had similar success rates (52.9% vs. 50%, P = 0.88). After failure of a first detrusor injection of botulinum toxin for neurogenic detrusor overactivity, a switch to a different toxin seems to be more effective than a second injection of the same toxin. The replacement of onabotulinumtoxin by abobotulinumtoxin or the reverse provides similar results. © 2015 The Japanese Urological Association.

Snake venomics of monocled cobra (Naja kaouthia) and investigation of human IgG response against venom toxins

DEFF Research Database (Denmark)

Laustsen, Andreas Hougaard; Gutiérrez, José María; Lohse, Brian

2015-01-01

/cardiotoxins. IgGs isolated from a person who had repeatedly self-immunized with a variety of snake venoms were immunoprofiled by ELISA against all venom fractions. Stronger responses against larger toxins, but lower against the most critical α-neurotoxins were obtained. As expected, no neutralization potential...

Bioinformatics and multiepitope DNA immunization to design rational snake antivenom.

Directory of Open Access Journals (Sweden)

Simon C Wagstaff

2006-06-01

Full Text Available Snake venom is a potentially lethal and complex mixture of hundreds of functionally diverse proteins that are difficult to purify and hence difficult to characterize. These difficulties have inhibited the development of toxin-targeted therapy, and conventional antivenom is still generated from the sera of horses or sheep immunized with whole venom. Although life-saving, antivenoms contain an immunoglobulin pool of unknown antigen specificity and known redundancy, which necessitates the delivery of large volumes of heterologous immunoglobulin to the envenomed victim, thus increasing the risk of anaphylactoid and serum sickness adverse effects. Here we exploit recent molecular sequence analysis and DNA immunization tools to design more rational toxin-targeted antivenom.We developed a novel bioinformatic strategy that identified sequences encoding immunogenic and structurally significant epitopes from an expressed sequence tag database of a venom gland cDNA library of Echis ocellatus, the most medically important viper in Africa. Focusing upon snake venom metalloproteinases (SVMPs that are responsible for the severe and frequently lethal hemorrhage in envenomed victims, we identified seven epitopes that we predicted would be represented in all isomers of this multimeric toxin and that we engineered into a single synthetic multiepitope DNA immunogen (epitope string. We compared the specificity and toxin-neutralizing efficacy of antiserum raised against the string to antisera raised against a single SVMP toxin (or domains or antiserum raised by conventional (whole venom immunization protocols. The SVMP string antiserum, as predicted in silico, contained antibody specificities to numerous SVMPs in E. ocellatus venom and venoms of several other African vipers. More significantly, the antiserum cross-specifically neutralized hemorrhage induced by E. ocellatus and Cerastes cerastes cerastes venoms.These data provide valuable sequence and structure

The central nervous system as target of Bacillus anthracis toxin independent virulence in rabbits and guinea pigs.

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Haim Levy

Full Text Available Infection of the central nervous system is considered a complication of Anthrax and was reported in humans and non-human primates. Previously we have reported that Bacillus anthracis possesses a toxin-independent virulent trait that, like the toxins, is regulated by the major virulence regulator, AtxA, in the presence of pXO2. This toxin-independent lethal trait is exhibited in rabbits and Guinea pigs following significant bacteremia and organ dissemination. Various findings, including meningitis seen in humans and primates, suggested that the CNS is a possible target for this AtxA-mediated activity. In order to penetrate into the brain tissue, the bacteria have to overcome the barriers isolating the CNS from the blood stream. Taking a systematic genetic approach, we compared intracranial (IC inoculation and IV/SC inoculation for the outcome of the infection in rabbits/GP, respectively. The outstanding difference between the two models is exhibited by the encapsulated strain VollumΔpXO1, which is lethal when injected IC, but asymptomatic when inoculated IV/SC. The findings demonstrate that there is an apparent bottleneck in the ability of mutants to penetrate into the brain. Any mutant carrying either pXO1 or pXO2 will kill the host upon IC injection, but only those carrying AtxA either on pXO1 or in the chromosome in the background of pXO2 can penetrate into the brain following peripheral inoculation. The findings were corroborated by histological examination by H&E staining and immunofluorescence of rabbits' brains following IV and IC inoculations. These findings may have major implications on future research both on B. anthracis pathogenicity and on vaccine development.

Theories of Lethal Mutagenesis: From Error Catastrophe to Lethal Defection.

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Tejero, Héctor; Montero, Francisco; Nuño, Juan Carlos

2016-01-01

RNA viruses get extinct in a process called lethal mutagenesis when subjected to an increase in their mutation rate, for instance, by the action of mutagenic drugs. Several approaches have been proposed to understand this phenomenon. The extinction of RNA viruses by increased mutational pressure was inspired by the concept of the error threshold. The now classic quasispecies model predicts the existence of a limit to the mutation rate beyond which the genetic information of the wild type could not be efficiently transmitted to the next generation. This limit was called the error threshold, and for mutation rates larger than this threshold, the quasispecies was said to enter into error catastrophe. This transition has been assumed to foster the extinction of the whole population. Alternative explanations of lethal mutagenesis have been proposed recently. In the first place, a distinction is made between the error threshold and the extinction threshold, the mutation rate beyond which a population gets extinct. Extinction is explained from the effect the mutation rate has, throughout the mutational load, on the reproductive ability of the whole population. Secondly, lethal defection takes also into account the effect of interactions within mutant spectra, which have been shown to be determinant for the understanding the extinction of RNA virus due to an augmented mutational pressure. Nonetheless, some relevant issues concerning lethal mutagenesis are not completely understood yet, as so survival of the flattest, i.e. the development of resistance to lethal mutagenesis by evolving towards mutationally more robust regions of sequence space, or sublethal mutagenesis, i.e., the increase of the mutation rate below the extinction threshold which may boost the adaptability of RNA virus, increasing their ability to develop resistance to drugs (including mutagens). A better design of antiviral therapies will still require an improvement of our knowledge about lethal

Recombinant Alpha, Beta, and Epsilon Toxins of Clostridium perfringens: Production Strategies and Applications as Veterinary Vaccines

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Marcos Roberto A. Ferreira

2016-11-01

Full Text Available Clostridium perfringens is a spore-forming, commensal, ubiquitous bacterium that is present in the gastrointestinal tract of healthy humans and animals. This bacterium produces up to 18 toxins. The species is classified into five toxinotypes (A–E according to the toxins that the bacterium produces: alpha, beta, epsilon, or iota. Each of these toxinotypes is associated with myriad different, frequently fatal, illnesses that affect a range of farm animals and humans. Alpha, beta, and epsilon toxins are the main causes of disease. Vaccinations that generate neutralizing antibodies are the most common prophylactic measures that are currently in use. These vaccines consist of toxoids that are obtained from C. perfringens cultures. Recombinant vaccines offer several advantages over conventional toxoids, especially in terms of the production process. As such, they are steadily gaining ground as a promising vaccination solution. This review discusses the main strategies that are currently used to produce recombinant vaccines containing alpha, beta, and epsilon toxins of C. perfringens, as well as the potential application of these molecules as vaccines for mammalian livestock animals.

A cocktail of humanized anti-pertussis toxin antibodies limits disease in murine and baboon models of whooping cough.

Science.gov (United States)

Nguyen, Annalee W; Wagner, Ellen K; Laber, Joshua R; Goodfield, Laura L; Smallridge, William E; Harvill, Eric T; Papin, James F; Wolf, Roman F; Padlan, Eduardo A; Bristol, Andy; Kaleko, Michael; Maynard, Jennifer A

2015-12-02

Despite widespread vaccination, pertussis rates are rising in industrialized countries and remain high worldwide. With no specific therapeutics to treat disease, pertussis continues to cause considerable infant morbidity and mortality. The pertussis toxin is a major contributor to disease, responsible for local and systemic effects including leukocytosis and immunosuppression. We humanized two murine monoclonal antibodies that neutralize pertussis toxin and expressed them as human immunoglobulin G1 molecules with no loss of affinity or in vitro neutralization activity. When administered prophylactically to mice as a binary cocktail, antibody treatment completely mitigated the Bordetella pertussis-induced rise in white blood cell counts and decreased bacterial colonization. When administered therapeutically to baboons, antibody-treated, but not untreated control animals, experienced a blunted rise in white blood cell counts and accelerated bacterial clearance rates. These preliminary findings support further investigation into the use of these antibodies to treat human neonatal pertussis in conjunction with antibiotics and supportive care. Copyright © 2015, American Association for the Advancement of Science.

Biliatresone, a Reactive Natural Toxin from Dysphania glomulifera and D. littoralis: Discovery of the Toxic Moiety 1,2-Diaryl-2-Propenone.

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Koo, Kyung A; Lorent, Kristin; Gong, Weilong; Windsor, Peter; Whittaker, Stephen J; Pack, Michael; Wells, Rebecca G; Porter, John R

2015-08-17

We identified a reactive natural toxin, biliatresone, from Dysphania glomulifera and D. littoralis collected in Australia that produces extrahepatic biliary atresia in a zebrafish model. Three additional isoflavonoids, including the known isoflavone betavulgarin, were also isolated. Biliatresone is in the very rare 1,2-diaryl-2-propenone class of isoflavonoids. The α-methylene of the 1,2-diaryl-2-propenone of biliatresone spontaneously reacts via Michael addition in the formation of water and methanol adducts. The lethal dose of biliatresone in a zebrafish assay was 1 μg/mL, while the lethal dose of synthetic 1,2-diaryl-2-propen-1-one was 5 μg/mL, suggesting 1,2-diaryl-2-propenone as the toxic Michael acceptor.

Bioterrorism: toxins as weapons.

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Anderson, Peter D

2012-04-01

The potential for biological weapons to be used in terrorism is a real possibility. Biological weapons include infectious agents and toxins. Toxins are poisons produced by living organisms. Toxins relevant to bioterrorism include ricin, botulinum, Clostridium perfrigens epsilson toxin, conotoxins, shigatoxins, saxitoxins, tetrodotoxins, mycotoxins, and nicotine. Toxins have properties of biological and chemical weapons. Unlike pathogens, toxins do not produce an infection. Ricin causes multiorgan toxicity by blocking protein synthesis. Botulinum blocks acetylcholine in the peripheral nervous system leading to muscle paralysis. Epsilon toxin damages cell membranes. Conotoxins block potassium and sodium channels in neurons. Shigatoxins inhibit protein synthesis and induce apoptosis. Saxitoxin and tetrodotoxin inhibit sodium channels in neurons. Mycotoxins include aflatoxins and trichothecenes. Aflatoxins are carcinogens. Trichothecenes inhibit protein and nucleic acid synthesis. Nicotine produces numerous nicotinic effects in the nervous system.

Toxicity of binary chemical munition destruction products: methylphosphonic acid, methylphosphinic acid, 2-diisopropylaminoethanol, DF neutralent, and QL neutralent.

Science.gov (United States)

Watson, Rebecca E; Hafez, Ahmed M; Kremsky, Jonathan N; Bizzigotti, George O

2007-01-01

This paper reports the toxicity and environmental impact of neutralents produced from the hydrolysis of binary chemical agent precursor chemicals DF (methylphosphonic difluoride) and QL (2-[bis(1-methylethyl)amino]ethyl ethyl methylphosphonite). Following a literature review of the neutralent mixtures and constituents, basic toxicity tests were conducted to fill data gaps, including acute oral and dermal median lethal dose assays, the Ames mutagenicity test, and ecotoxicity tests. For methylphosphonic acid (MPA), a major constituent of DF neutralent, the acute oral LD(50) in the Sprague-Dawley rat was measured at 1888 mg/kg, and the Ames test using typical tester strains of Salmonella typhimurium and Escherichia coli was negative. The 48-h LC(50) values for pH-adjusted DF neutralent with Daphnia magna and Cyprinodon variegatus were > 2500 mg/L and 1593 mg/L, respectively. The acute oral LD(50) values in the rat for QL neutralent constituents methylphosphinic acid (MP) and 2-diisopropylaminoethanol (KB) were both determined to be 940 mg/kg, and the Ames test was negative for both. Good Laboratory Practice (GLP)-compliant ecotoxicity tests for MP and KB gave 48-h D. magna EC(50) values of 6.8 mg/L and 83 mg/L, respectively. GLP-compliant 96-h C. variegatus assays on MP and KB gave LC(50) values of 73 and 252 mg/L, respectively, and NOEC values of 22 and 108 mg/L. QL neutralent LD(50) values for acute oral and dermal toxicity tests were both > 5000 mg/kg, and the 48-h LD(50) values for D. magna and C. variegatus were 249 and 2500 mg/L, respectively. Using these data, the overall toxicity of the neutralents was assessed.

Environmental T4-Family Bacteriophages Evolve to Escape Abortive Infection via Multiple Routes in a Bacterial Host Employing "Altruistic Suicide" through Type III Toxin-Antitoxin Systems.

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Chen, Bihe; Akusobi, Chidiebere; Fang, Xinzhe; Salmond, George P C

2017-01-01

Abortive infection is an anti-phage mechanism employed by a bacterium to initiate its own death upon phage infection. This reduces, or eliminates, production of viral progeny and protects clonal siblings in the bacterial population by an act akin to an "altruistic suicide." Abortive infection can be mediated by a Type III toxin-antitoxin system called ToxIN Pa consisting of an endoribonuclease toxin and RNA antitoxin. ToxIN Pa is a heterohexameric quaternary complex in which pseudoknotted RNA inhibits the toxicity of the toxin until infection by certain phages causes destabilization of ToxIN Pa , leading to bacteriostasis and, eventually, lethality. However, it is still unknown why only certain phages are able to activate ToxIN Pa . To try to address this issue we first introduced ToxIN Pa into the Gram-negative enterobacterium, Serratia sp. ATCC 39006 ( S 39006) and then isolated new environmental S 39006 phages that were scored for activation of ToxIN Pa and abortive infection capacity. We isolated three T4-like phages from a sewage treatment outflow point into the River Cam, each phage being isolated at least a year apart. These phages were susceptible to ToxIN Pa -mediated abortive infection but produced spontaneous "escape" mutants that were insensitive to ToxIN Pa . Analysis of these resistant mutants revealed three different routes of escaping ToxIN Pa , namely by mutating asiA (the product of which is a phage transcriptional co-activator); by mutating a conserved, yet functionally unknown, orf84 ; or by deleting a 6.5-10 kb region of the phage genome. Analysis of these evolved escape mutants may help uncover the nature of the corresponding phage product(s) involved in activation of ToxIN Pa .

Suicide Lethality: A Concept Analysis.

Science.gov (United States)

DeBastiani, Summer; De Santis, Joseph P

2018-02-01

Suicide is a significant health problem internationally. Those who complete suicide may have different behaviors and risk factors than those who attempt a non-fatal suicide. The purpose of this article is to analyze the concept of suicide lethality and propose a clear definition of the concept through the identification of antecedents, attributes, and consequences. A literature search for articles published in the English language between 1970 and 2016 was conducted using MEDLINE, the Cochrane Library, Pubmed, Psychlit, Ovid, PsycINFO, and Proquest. The bibliographies of all included studies were also reviewed to identify additional relevant citations. A concept analysis was conducted on the literature findings using six stages of Walker and Avant's method. The concept analysis differentiated between suicide, lethality, suicidal behavior, and suicide lethality. Presence of a suicide plan or a written suicide note was not found to be associated with the majority of completed suicides included in the definition of suicide lethality. There are a few scales that measure the lethality of a suicide attempt, but none that attempt to measure the concept of suicide lethality as described in this analysis. Clarifying the concept of suicide lethality encourages awareness of the possibility of different suicidal behaviors associated with different suicide outcomes and will inform the development of future nursing interventions. A clearer definition of the concept of suicide lethality will guide clinical practice, research, and policy development aimed at suicide prevention.

Botulinum toxin

Directory of Open Access Journals (Sweden)

Nigam P

2010-01-01

Full Text Available Botulinum toxin, one of the most poisonous biological substances known, is a neurotoxin produced by the bacterium Clostridium botulinum. C. botulinum elaborates eight antigenically distinguishable exotoxins (A, B, C 1 , C 2 , D, E, F and G. All serotypes interfere with neural transmission by blocking the release of acetylcholine, the principal neurotransmitter at the neuromuscular junction, causing muscle paralysis. The weakness induced by injection with botulinum toxin A usually lasts about three months. Botulinum toxins now play a very significant role in the management of a wide variety of medical conditions, especially strabismus and focal dystonias, hemifacial spasm, and various spastic movement disorders, headaches, hypersalivation, hyperhidrosis, and some chronic conditions that respond only partially to medical treatment. The list of possible new indications is rapidly expanding. The cosmetological applications include correction of lines, creases and wrinkling all over the face, chin, neck, and chest to dermatological applications such as hyperhidrosis. Injections with botulinum toxin are generally well tolerated and side effects are few. A precise knowledge and understanding of the functional anatomy of the mimetic muscles is absolutely necessary to correctly use botulinum toxins in clinical practice.

Identification of the factors that govern the ability of therapeutic antibodies to provide postchallenge protection against botulinum toxin: a model for assessing postchallenge efficacy of medical countermeasures against agents of bioterrorism and biological warfare.

Science.gov (United States)

Al-Saleem, Fetweh H; Nasser, Zidoon; Olson, Rebecca M; Cao, Linsen; Simpson, Lance L

2011-08-01

Therapeutic antibodies are one of the major classes of medical countermeasures that can provide protection against potential bioweapons such as botulinum toxin. Although a broad array of antibodies are being evaluated for their ability to neutralize the toxin, there is little information that defines the circumstances under which these antibodies can be used. In the present study, an effort was made to quantify the temporal factors that govern therapeutic antibody use in a postchallenge scenario. Experiments were done involving inhalation administration of toxin to mice, intravenous administration to mice, and direct application to murine phrenic nerve-hemidiaphragm preparations. As part of this study, several pharmacokinetic characteristics of botulinum toxin and neutralizing antibodies were measured. The core observation that emerged from the work was that the window of opportunity within which postchallenge administration of antibodies exerted a beneficial effect increased as the challenge dose of toxin decreased. The critical factor in establishing the window of opportunity was the amount of time needed for fractional redistribution of a neuroparalytic quantum of toxin from the extraneuronal space to the intraneuronal space. This redistribution event was a dose-dependent phenomenon. It is likely that the approach used to identify the factors that govern postchallenge efficacy of antibodies against botulinum toxin can be used to assess the factors that govern postchallenge efficacy of medical countermeasures against any agent of bioterrorism or biological warfare.

The Intrinsically Disordered Domain of the Antitoxin Phd Chaperones the Toxin Doc against Irreversible Inactivation and Misfolding*

Science.gov (United States)

De Gieter, Steven; Konijnenberg, Albert; Talavera, Ariel; Butterer, Annika; Haesaerts, Sarah; De Greve, Henri; Sobott, Frank; Loris, Remy; Garcia-Pino, Abel

2014-01-01

The toxin Doc from the phd/doc toxin-antitoxin module targets the cellular translation machinery and is inhibited by its antitoxin partner Phd. Here we show that Phd also functions as a chaperone, keeping Doc in an active, correctly folded conformation. In the absence of Phd, Doc exists in a relatively expanded state that is prone to dimerization through domain swapping with its active site loop acting as hinge region. The domain-swapped dimer is not capable of arresting protein synthesis in vitro, whereas the Doc monomer is. Upon binding to Phd, Doc becomes more compact and is secured in its monomeric state with a neutralized active site. PMID:25326388

Botulinum toxin in parkinsonism: The when, how, and which for botulinum toxin injections.

Science.gov (United States)

Cardoso, Francisco

2018-06-01

The aim of this article is to provide a review of the use of injections of botulinum toxin in the management of selected symptoms and signs of Parkinson's disease and other forms of parkinsonism. Sialorrhea is defined as inability to control oral secretions, resulting in excessive saliva in the oropharynx. There is a high level of evidence for the treatment of sialorrhea in parkinsonism with injections of different forms of botulinum toxin type A as well as botulinum toxin type B. Tremor can be improved by the use of botulinum toxin injections but improved tremor control often leads to concomitant motor weakness, limiting its use. Levodopa induced dyskinesias are difficult to treat with botulinum toxin injections because of their variable frequency and direction. Apraxia of eyelid opening, a sign more commonly seen in progressive supranuclear palsy and other tauopathies, often improves after botulinum toxin injections. Recent data suggest that regardless of the underlying mechanism, pain in parkinsonism can be alleviated by botulinum toxin injections. Finally, freezing of gait, camptocormia and Pisa syndrome in parkinsonism almost invariably fail to respond to botulinum toxin injections. Copyright © 2017 Elsevier Ltd. All rights reserved.

Occurrence of tetrodotoxin-binding high molecular weight substances in the body fluid of shore crab (Hemigrapsus sanguineus).

Science.gov (United States)

Shiomi, K; Yamaguchi, S; Kikuchi, T; Yamamori, K; Matsui, T

1992-12-01

The shore crab (Hemigrapsus sanguineus) is highly resistant to tetrodotoxin (TTX) although it contains no detectable amount of TTX (less than 5 MU/g, where 1 MU is defined as the amount of TTX killing a 20 g mouse in 30 min). Its body fluid was examined for neutralizing effects against the lethal activity of TTX. When the mixture of the body fluid and TTX was injected i.p. into mice, the lethal activity of TTX was significantly reduced; 1 ml of the body fluid was evaluated to neutralize 3.6-4.0 MU of TTX. Higher neutralizing activity (7.2-12.5 MU/ml of the body fluid) was exhibited by i.v. administration of the body fluid into mice before or after i.p. challenge of TTX. The lethal effect of paralytic shellfish poisons was not counteracted by the body fluid. Analysis by gel filtration on Sepharose 6B revealed that the body fluid contained TTX-binding high mol. wt substances (> 2,000,000) responsible for the neutralizing activity of the body fluid against TTX, which accounts for the high resistibility of the crab to TTX. When the crude toxin extracted from the liver of puffer (Takifugu niphobles) was mixed with the body fluid and chromatographed on Sepharose 6B, almost pure TTX was obtained from the fractions containing the TTX-binding high mol. wt substances, suggesting that the TTX-binding high mol. wt substances could be useful in purification of TTX from biological samples.

Snake venom neutralization by Indian medicinal plants (Vitex negundo and Emblica officinalis) root extracts.

Science.gov (United States)

Alam, M I; Gomes, A

2003-05-01

The methanolic root extracts of Vitex negundo Linn. and Emblica officinalis Gaertn. were explored for the first time for antisnake venom activity. The plant (V. negundo and E. officinalis) extracts significantly antagonized the Vipera russellii and Naja kaouthia venom induced lethal activity both in in vitro and in vivo studies. V. russellii venom-induced haemorrhage, coagulant, defibrinogenating and inflammatory activity was significantly neutralized by both plant extracts. No precipitating bands were observed between the plant extract and snake venom. The above observations confirmed that the plant extracts possess potent snake venom neutralizing capacity and need further investigation.

Polyamine toxins

DEFF Research Database (Denmark)

Strømgaard, Kristian; Jensen, Lars S; Vogensen, Stine B

2005-01-01

Polyamine toxins, isolated from spiders and wasps, have been used as pharmacological tools for the study of ionotropic receptors, but their use have so far been hampered by their lack of selectivity. In this mini-review, we describe how careful synthetic modification of native polyamine toxins ha...

Action of cholera toxin in the intestinal epithelial cells

International Nuclear Information System (INIS)

Hyun, C.S.

1982-01-01

The primary event in the action of cholera toxin on the isolated chick intestinal epithelial cell is its interaction with a large number of high affinity binding sites in the cell membrane. Binding of 125 I-labeled toxin is rapid, temperature-dependent, reversible, and saturable over a wide range of concentrations and includes only a small contribution from nonspecific sites. A characteristic lag phase of 10 min occurs following the complete binding of toxin before any increase in cellular cAMP levels can be detected. The response (elevation of cellular cAMP) is linear with time for 40 to 50 min and causes a six- to eight-fold increase over control levels (10 to 15 picomole cAMP/mg cellular protein) at steady state. cAMP and agents that increase cAMP production inhibit Cl - -independent Na + influx into the isolated enterocytes whereas chlorpromazine (CPZ) which completely abolishes toxin-induced elevation of cAMP both reverses and prevents the cAMP-mediated inhibition of Na + entry. Correlation between cellular cAMP levels and the magnitude of Na + influx provides evidence for a cAMP-mediated control of intestinal Na + uptake, which may represent the mechanistic basis for the antiabsorptive effect of CT on Na + during induction of intestinal secretion. The effect of cAMP on Na + but not Cl - influx preparations can be partially explained in terms of a cAMP-regulated Na + /H + neutral exchange system. Data on the coupling relationship between Na + transport and the intra- and extracellular pH in the enterocytes show that an amiloride-sensitive electroneutral Na + /H + exchange process occurs. This coupling between Na + and H + is partially inhibited by CT and dbcAMP, suggesting that the Na + /H + exchange may be a cAMP-regulated process. 31 references, 32 figures, 5 tables

Higher cytotoxicity of divalent antibody-toxins than monovalent antibody-toxins

International Nuclear Information System (INIS)

Won, JaeSeon; Nam, PilWon; Lee, YongChan; Choe, MuHyeon

2009-01-01

Recombinant antibody-toxins are constructed via the fusion of a 'carcinoma-specific' antibody fragment to a toxin. Due to the high affinity and high selectivity of the antibody fragments, antibody-toxins can bind to surface antigens on cancer cells and kill them without harming normal cells [L.H. Pai, J.K. Batra, D.J. FitzGerald, M.C. Willingham, I. Pastan, Anti-tumor activities of immunotoxins made of monoclonal antibody B3 and various forms of Pseudomonas exotoxin, Proc. Natl. Acad. Sci. USA 88 (1991) 3358-3362]. In this study, we constructed the antibody-toxin, Fab-SWn-PE38, with SWn (n = 3, 6, 9) sequences containing n-time repeated (G 4 S) between the Fab fragment and PE38 (38 kDa truncated form of Pseudomonas exotoxin A). The SWn sequence also harbored one cysteine residue that could form a disulfide bridge between two Fab-SWn-PE38 monomers. We assessed the cytotoxicity of the monovalent (Fab-SWn-PE38), and divalent ([Fab-SWn-PE38] 2 ) antibody-toxins. The cytotoxicity of the dimer against the CRL1739 cell line was approximately 18.8-fold higher than that of the monomer on the ng/ml scale, which was approximately 37.6-fold higher on the pM scale. These results strongly indicate that divalency provides higher cytotoxicity for an antibody-toxin.

Quiescent complement in nonhuman primates during E coli Shiga toxin-induced hemolytic uremic syndrome and thrombotic microangiopathy.

Science.gov (United States)

Lee, Benjamin C; Mayer, Chad L; Leibowitz, Caitlin S; Stearns-Kurosawa, D J; Kurosawa, Shinichiro

2013-08-01

Enterohemorrhagic Escherichia coli (EHEC) produce ribosome-inactivating Shiga toxins (Stx1, Stx2) responsible for development of hemolytic uremic syndrome (HUS) and acute kidney injury (AKI). Some patients show complement activation during EHEC infection, raising the possibility of therapeutic targeting of complement for relief. Our juvenile nonhuman primate (Papio baboons) models of endotoxin-free Stx challenge exhibit full spectrum HUS, including thrombocytopenia, hemolytic anemia, and AKI with glomerular thrombotic microangiopathy. There were no significant increases in soluble terminal complement complex (C5b-9) levels after challenge with lethal Stx1 (n = 6) or Stx2 (n = 5) in plasma samples from T0 to euthanasia at 49.5 to 128 hours post-challenge. d-dimer and cell injury markers (HMGB1, histones) confirmed coagulopathy and cell injury. Thus, complement activation is not required for the development of thrombotic microangiopathy and HUS induced by EHEC Shiga toxins in these preclinical models, and benefits or risks of complement inhibition should be studied further for this infection.

Structural Basis for the Binding of the Neutralizing Antibody, 7D11, to the Poxvirus L1 Protein

Science.gov (United States)

2007-08-01

pCR- 7D11-vHC and pCR-7D11- vLC , respectively. Crystallization of the complex between L1 and 7D11-Fab VACV L1 protein was expressed and purified as...2005. Vaccinia virus H3L envelope protein is a major target of neutralizing antibodies in humans and elicits protection against lethal challenge in...D.M., Schmaljohn, C., Schmaljohn, A., 2000. DNA vaccination with vaccinia virus L1R and A33R genes protects mice against a lethal poxvirus challenge

Specificity of antibodies directed against the cytolethal distending toxin of Haemophilus ducreyi in patients with chancroid.

Science.gov (United States)

Mbwana, Judica; Ahmed, Hinda J; Ahlman, Karin; Sundaeus, Vivian; Dahlén, Gunnar; Lyamuya, Eligius; Lagergård, Teresa

2003-09-01

Antibodies specific for the cytolethal-distending toxin of Haemophilus ducreyi (HdCDT) complex and for the CdtA, CdtB, and CdtC components were measured by ELISA in the sera of 50 patients with culture and/or PCR proven chancroid, 42 patients with periodontitis, 50 blood donors from Tanzania, 50 blood donors from Sweden. In addition, the biological activity e.g. neutralization capacity of the sera were tested. Our results demonstrate that majority of chancroid patients and healthy individuals had detectable levels of serum antibodies to HdCDT complex and to separate toxin components. However, high levels (> or =100 units) of antibodies to HdCDT complex were significantly more prevalent in the sera of patients with both chancroid and periodontitis than in the sera of the corresponding controls (P=0.001 and P=0.04, respectively). In the sera of the 50 patients with chancroid, antibodies to CdtA, CdtB, and CdtC were detected in 50, 35, and 34 individuals, respectively. Antibodies to CdtC, being less frequently detected than the antibodies to other components, show a good correlation with the neutralizing capacity of sera. High levels of neutralizing antibodies (> or =160) were detected in only 22 and 2% of the patients with chancroid and periodontitis, respectively. The data suggest that the low levels of anti-HdCDT antibodies, which include neutralizing antibodies, may contribute to limited protection in chancroid and since anti-HdCDT antibodies, may be detected in healthy individuals and in patients with certain disease conditions (e.g. periodontitis), they may not be specific markers for chancroid infection.

Authentic display of a cholera toxin epitope by chimeric type 1 fimbriae: effects of insert position and host background

DEFF Research Database (Denmark)

Stentebjerg-Olesen, B; Pallesen, L; Jensen, LB

1997-01-01

The potential of the major structural protein of type 1 fimbriae as a display system for heterologous sequences was tested. As a reporter-epitope, a heterologous sequence mimicking a neutralizing epitope of the cholera toxin B chain was inserted, in one or two copies, into four different positions...... in the fimA gene. This was carried out by introduction of new restriction sites by PCR-mediated site-directed mutagenesis of fimA in positions predicted to correspond to optimally surface-located regions of the subunit protein. Subsequently, the synthetic cholera-toxin-encoding DNA segment was inserted....... Several of the chosen positions seemed amenable even for large foreign inserts; the chimeric proteins were exposed on the bacterial surface and the cholera toxin epitope was authentically displayed, i.e. it was recognized on bacteria by specific antiserum. Display of chimeric fimbriae was tested...

Tumor therapy with a urokinase plasminogen activator-activated anthrax lethal toxin alone and in combination with paclitaxel

OpenAIRE

Wein, Alexander N.; Liu, Shihui; Zhang, Yi; McKenzie, Andrew T.; Leppla, Stephen H.

2012-01-01

PA-U2, an engineered anthrax protective antigen that is activated by urokinase was combined with wild-type lethal factor in the treatment of Colo205 colon adenocarcinoma in vitro and B16-BL6 mouse melanoma in vitro and in vivo. This therapy was also tested in combination with the small molecule paclitaxel, based on prior reports suggesting synergy between ERK1/2 inhibition and chemotherapeutics. Colo205 was sensitive to PA-U2/LF while B16-BL6 was not. For the combination treatment of B16-BL6,...

Comparison of Phylogeny, Venom Composition and Neutralization by Antivenom in Diverse Species of Bothrops Complex

Science.gov (United States)

Peixoto, Pedro S.; Bernardoni, Juliana L.; Oliveira, Sâmella S.; Portes-Junior, José Antonio; Mourão, Rosa Helena V.; Lima-dos-Santos, Isa; Sano-Martins, Ida S.; Chalkidis, Hipócrates M.; Valente, Richard H.; Moura-da-Silva, Ana M.

2013-01-01

In Latin America, Bothrops snakes account for most snake bites in humans, and the recommended treatment is administration of multispecific Bothrops antivenom (SAB – soro antibotrópico). However, Bothrops snakes are very diverse with regard to their venom composition, which raises the issue of which venoms should be used as immunizing antigens for the production of pan-specific Bothrops antivenoms. In this study, we simultaneously compared the composition and reactivity with SAB of venoms collected from six species of snakes, distributed in pairs from three distinct phylogenetic clades: Bothrops, Bothropoides and Rhinocerophis. We also evaluated the neutralization of Bothrops atrox venom, which is the species responsible for most snake bites in the Amazon region, but not included in the immunization antigen mixture used to produce SAB. Using mass spectrometric and chromatographic approaches, we observed a lack of similarity in protein composition between the venoms from closely related snakes and a high similarity between the venoms of phylogenetically more distant snakes, suggesting little connection between taxonomic position and venom composition. P-III snake venom metalloproteinases (SVMPs) are the most antigenic toxins in the venoms of snakes from the Bothrops complex, whereas class P-I SVMPs, snake venom serine proteinases and phospholipases A2 reacted with antibodies in lower levels. Low molecular size toxins, such as disintegrins and bradykinin-potentiating peptides, were poorly antigenic. Toxins from the same protein family showed antigenic cross-reactivity among venoms from different species; SAB was efficient in neutralizing the B. atrox venom major toxins. Thus, we suggest that it is possible to obtain pan-specific effective antivenoms for Bothrops envenomations through immunization with venoms from only a few species of snakes, if these venoms contain protein classes that are representative of all species to which the antivenom is targeted. PMID

Autoproteolytic Activation of Bacterial Toxins

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Aimee Shen

2010-05-01

Full Text Available Protease domains within toxins typically act as the primary effector domain within target cells. By contrast, the primary function of the cysteine protease domain (CPD in Multifunctional Autoprocessing RTX-like (MARTX and Clostridium sp. glucosylating toxin families is to proteolytically cleave the toxin and release its cognate effector domains. The CPD becomes activated upon binding to the eukaryotic-specific small molecule, inositol hexakisphosphate (InsP6, which is found abundantly in the eukaryotic cytosol. This property allows the CPD to spatially and temporally regulate toxin activation, making it a prime candidate for developing anti-toxin therapeutics. In this review, we summarize recent findings related to defining the regulation of toxin function by the CPD and the development of inhibitors to prevent CPD-mediated activation of bacterial toxins.

Toxin production in Dinophysis and the fate of these toxins in marine mussels

DEFF Research Database (Denmark)

Nielsen, Lasse Tor

Diarrhetic shellfish poisoning (DSP) poses a considerable threat to food safety and to the economy of shellfish fishers and farmers in many parts of the world. Thousands of DSP intoxications have been reported, and bivalve harvesting can sometimes be closed down several months in a row. The toxins....... acuta. I grew the two species in laboratory cultures at different irradiances (7-130 μmol photons m-2 s-1) and with different food availability. The results showed that irradiance had no effects on toxin profiles, and only limited effects of the cellular toxin contents. Rather, toxin production rates...... are primarily produced by the marine mixotrophic dinoflagellates Dinophysis spp., known to occur in most parts of the world. Dinophysis can, along with other planktonic organisms, be consumed by filter-feeding bivalves, and thus the toxins can accumulate. Dinophysis can produce the three toxin groups, okadaic...

Immunoglobulin G and F(ab')2 polyvalent antivenoms do not differ in their ability to neutralize hemorrhage, edema and myonecrosis induced by Bothrops asper (terciopelo) snake venom

OpenAIRE

León Montero, Guillermo; Rojas Céspedes, Gustavo; Lomonte, Bruno; Gutiérrez, José María

1997-01-01

The ability of whole immunoglobulin G (IgG) and F(ab')2 polyvalent (Crotalinae) antivenoms to neutralize the hemorrhagic, edema-forming and myotoxic activities of Bothrops asper venom was studied. Both antivenoms were adjusted to the same neutralizing potency against lethal and hemorrhagic activities in experiments where venom and antivenoms were incubated before injection. Thus, in these experimental conditions, differences in the neutralizing ability in experiments involving independent inj...

Affinity chromatography of tetanus toxin, tetanus toxoid, and botulinum A toxin on synaptosomes, and differentiation of their acceptors

Energy Technology Data Exchange (ETDEWEB)

Habermann, E [Giessen Univ. (Germany, F.R.). Pharmakologisches Inst.

1976-01-01

/sup 125/I-labelled tetanus toxin and /sup 125/I-labelled botulinum A neurotoxin are known to be specifically bound to brain synaptosomes. In order to discriminate between active toxin and inactive admixtures present in the starting material or arising during iodination, synaptosome columns were prepared using bromacetylcellulose and/or kieselgur (Celite) as carriers. Both types of columns adsorb the toxins from low ionic strength medium and release them if the pH and ionic strength are raised. Botulinum toxin was eluted with lower ionic strength than tetanus toxin, and could be freed from nontoxic admixtures. Analysis by affinity chromatography disclosed partially toxoided tetanus toxin in both labelled and unlabelled toxin samples. High concentrations of formaldehyde (0.5%) destroyed both toxicity and affinity to the synaptosomes of tetanus toxin. Low concentrations of formaldehyde (0.05%) yielded a derivative of low toxicity which was still, however less firmly, bound to synaptosomes. Tetanus and botulinum toxin differ by their acceptors. Whereas unlabelled botulinum toxin is unable to compete with labelled tetanus toxin, unlabelled tetanus toxin slightly competes with botulinum toxin. Both labelled toxins display anomalous binding behaviour in that they cannot be displaced completely even with a large excess of unlabelled toxin.

Affinity chromatography of tetanus toxin, tetanus toxoid, and botulinum A toxin on synaptosomes, and differentiation of their acceptors

International Nuclear Information System (INIS)

Habermann, E.

1976-01-01

125 I-labelled tetanus toxin and 125 I-labelled botulinum A neurotoxin are known to be specifically bound to brain synaptosomes. In order to discriminate between active toxin and inactive admixtures present in the starting material or arising during iodination, synaptosome columns were prepared using bromacetylcellulose and/or kieselgur (Celite) as carriers. Both types of columns adsorb the toxins from low ionic strength medium and release them if the pH and ionic strength are raised. Botulinum toxin was eluted with lower ionic strength than tetanus toxin, and could be freed from nontoxic admixtures. Analysis by affinity chromatography disclosed partially toxoided tetanus toxin in both labelled and unlabelled toxin samples. High concentrations of formaldehyde (0.5%) destroyed both toxicity and affinity to the synaptosomes of tetanus toxin. Low concentrations of formaldehyde (0.05%) yielded a derivative of low toxicity which was still, however less firmly, bound to synaptosomes. Tetanus and botulinum toxin differ by their acceptors. Whereas unlabelled botulinum toxin is unable to compete with labelled tetanus toxin, unlabelled tetanus toxin slightly competes with botulinum toxin. Both labelled toxins display anomalous binding behaviour in that they cannot be displaced completely even with a large excess of unlabelled toxin. (orig.) [de

Immunization of pregnant cows with Shiga toxin-2 induces high levels of specific colostral antibodies and lactoferrin able to neutralize E. coli O157:H7 pathogenicity.

Science.gov (United States)

Albanese, Adriana; Sacerdoti, Flavia; Seyahian, E Abril; Amaral, Maria Marta; Fiorentino, Gabriela; Fernandez Brando, Romina; Vilte, Daniel A; Mercado, Elsa C; Palermo, Marina S; Cataldi, Angel; Zotta, Elsa; Ibarra, Cristina

2018-03-20

E. coli O157:H7 is a foodborne pathogen responsible for bloody diarrhea, hemorrhagic colitis and hemolytic uremic syndrome (HUS). The objective of the present work was to evaluate the ability of colostral IgG obtained from Stx2-immunized cows to prevent against E. coli O157:H7 infection and Stx2 cytotoxicity. Hyperimmune colostrum (HC) was obtained from cows intramuscularly immunized with inactivated Stx2 or vehicle for controls. Colostral IgG was purified by affinity chromatography. Specific IgG antibodies against Stx2 and bovine lactoferrin (bLF) levels in HC and the corresponding IgG (HC-IgG/bLF) were determined by ELISA. The protective effects of HC-IgG/bLF against Stx2 cytotoxicity and adhesion of E. coli O157:H7 and its Stx2-negative mutant were analyzed in HCT-8 cells. HC-IgG/bLF prevention against E. coli O157:H7 was studied in human colon and rat colon loops. Protection against a lethal dose of E. coli O157:H7 was evaluated in a weaned mice model. HC-IgG/bLF showed high anti-Stx2 titers and high bLF levels that were able to neutralize the cytotoxic effects of Stx2 in vitro and in vivo. Furthermore, HC-IgG/bLF avoided the inhibition of water absorption induced by E. coli O157:H7 in human colon and also the pathogenicity of E. coli O157:H7 and E. coli O157:H7Δstx2 in rat colon loops. Finally, HC-IgG/bLF prevented in a 100% the lethality caused by E. coli O157:H7 in a weaned mice model. Our study suggests that HC-IgG/bLF have protective effects against E. coli O157:H7 infection. These beneficial effects may be due to specific anti-Stx2 neutralizing antibodies in combination with high bLF levels. These results allow us to consider HC-IgG/bLF as a nutraceutical tool which could be used in combination with balanced supportive diets to prevent HUS. However further studies are required before recommendations can be made for therapeutic and clinical applications. Copyright © 2018 Elsevier Ltd. All rights reserved.

Diabetes susceptibility of BALB/cBOM mice treated with streptozotocin. Inhibition by lethal irradiation and restoration by splenic lymphocytes

International Nuclear Information System (INIS)

Paik, S.G.; Blue, M.L.; Fleischer, N.; Shin, S.

1982-01-01

In genetically susceptible strains of mice, repeated injections of a subdiabetogenic dose of streptozotocin induces the development of progressive insulin-dependent hyperglycemia. We showed previously that host T-cell functions play an obligatory etiologic role in this experimental disease by demonstrating that the athymic nude mouse is resistant to diabetes induction unless its T-cell functions are reconstituted by thymus graft. Here we show that lethal irradiation of euthymic (+/nu) mice of BALB/cBOM background causes selective resistance of the mice to the diabetogenic effects of the multiple low doses of streptozotocin without affecting their sensitivity to a high pharmacologic dose of the toxin. We also show that reconstitution of the irradiated mice with splenic lymphocytes causes the restoration of diabetes susceptibility. Lethally irradiated mice thus represent a useful experimental model for analyzing the host functions involved in the development of this disease. These results provide an additional support for the hypothesis that the induction of diabetes in this model system is mediated by an autoimmune amplification mechanism

Clostridium Perfringens Epsilon Toxin Binds to Membrane Lipids and Its Cytotoxic Action Depends on Sulfatide.

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Carles Gil

Full Text Available Epsilon toxin (Etx is one of the major lethal toxins produced by Clostridium perfringens types B and D, being the causal agent of fatal enterotoxemia in animals, mainly sheep and goats. Etx is synthesized as a non-active prototoxin form (proEtx that becomes active upon proteolytic activation. Etx exhibits a cytotoxic effect through the formation of a pore in the plasma membrane of selected cell targets where Etx specifically binds due to the presence of specific receptors. However, the identity and nature of host receptors of Etx remain a matter of controversy. In the present study, the interactions between Etx and membrane lipids from the synaptosome-enriched fraction from rat brain (P2 fraction and MDCK cell plasma membrane preparations were analyzed. Our findings show that both Etx and proEtx bind to lipids extracted from lipid rafts from the two different models as assessed by protein-lipid overlay assay. Lipid rafts are membrane microdomains enriched in cholesterol and sphingolipids. Binding of proEtx to sulfatide, phosphatidylserine, phosphatidylinositol (3-phosphate and phosphatidylinositol (5-phosphate was detected. Removal of the sulphate groups via sulfatase treatment led to a dramatic decrease in Etx-induced cytotoxicity, but not in proEtx-GFP binding to MDCK cells or a significant shift in oligomer formation, pointing to a role of sulfatide in pore formation in rafts but not in toxin binding to the target cell membrane. These results show for the first time the interaction between Etx and membrane lipids from host tissue and point to a major role for sulfatides in C. perfringens epsilon toxin pathophysiology.

Acute Oral Toxicity of Tetrodotoxin in Mice: Determination of Lethal Dose 50 (LD50 and No Observed Adverse Effect Level (NOAEL

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Paula Abal

2017-02-01

Full Text Available Tetrodotoxin (TTX is starting to appear in molluscs from the European waters and is a hazard to seafood consumers. This toxin blocks sodium channels resulting in neuromuscular paralysis and even death. As a part of the risk assessment process leading to a safe seafood level for TTX, oral toxicity data are required. In this study, a 4-level Up and Down Procedure was designed in order to determine for the first time the oral lethal dose 50 (LD50 and the No Observed Adverse Effect Level (NOAEL in mice by using an accurate well-characterized TTX standard.

Comparison of toxicity neutralization-, ELISA- and PCR tests for typing of Clostridium perfringens and detection of the enterotoxin gene by PCR

DEFF Research Database (Denmark)

Møller, Kristian; Ahrens, Peter

1996-01-01

A polymerase chain reaction (PCR) was developed for the specific amplification of a part of each of the five Clostridium perfringens toxin genes: alpha (alpha), beta (beta), epsilon (epsilon), iota (iota), and enterotoxin (CPE). While the toxicity neutralization test (TNT) only showed limited...

Exploration of immunoglobulin transcriptomes from mice immunized with three-finger toxins and phospholipases A2 from the Central American coral snake, Micrurus nigrocinctus

DEFF Research Database (Denmark)

Laustsen, Andreas Hougaard; Engmark, Mikael; Clouser, Christopher

2017-01-01

Snakebite envenomings represent a neglected public health issue in many parts of the rural tropical world. Animal-derived antivenoms have existed for more than a hundred years and are effective in neutralizing snake venom toxins when timely administered. However, the low immunogenicity of many...

The intrinsically disordered domain of the antitoxin Phd chaperones the toxin Doc against irreversible inactivation and misfolding.

Science.gov (United States)

De Gieter, Steven; Konijnenberg, Albert; Talavera, Ariel; Butterer, Annika; Haesaerts, Sarah; De Greve, Henri; Sobott, Frank; Loris, Remy; Garcia-Pino, Abel

2014-12-05

The toxin Doc from the phd/doc toxin-antitoxin module targets the cellular translation machinery and is inhibited by its antitoxin partner Phd. Here we show that Phd also functions as a chaperone, keeping Doc in an active, correctly folded conformation. In the absence of Phd, Doc exists in a relatively expanded state that is prone to dimerization through domain swapping with its active site loop acting as hinge region. The domain-swapped dimer is not capable of arresting protein synthesis in vitro, whereas the Doc monomer is. Upon binding to Phd, Doc becomes more compact and is secured in its monomeric state with a neutralized active site. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.

Treatment of an Aedes aegypti colony with the Cry11Aa toxin for 54 generations results in the development of resistance

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Gloria Cadavid-Restrepo

2012-02-01

Full Text Available To study the potential for the emergence of resistance in Aedes aegypti populations, a wild colony was subjected to selective pressure with Cry11Aa, one of four endotoxins that compose the Bacillus thuringiensis serovar israelensis toxin. This bacterium is the base component of the most important biopesticide used in the control of mosquitoes worldwide. After 54 generations of selection, significant resistance levels were observed. At the beginning of the selection experiment, the half lethal concentration was 26.3 ng/mL and had risen to 345.6 ng/mL by generation 54. The highest rate of resistance, 13.1, was detected in the 54th generation. Because digestive proteases play a key role in the processing and activation of B. thuringiensis toxin, we analysed the involvement of insect gut proteases in resistance to the Cry11Aa B. thuringiensis serovar israelensis toxin. The protease activity from larval gut extracts from the Cry11Aa resistant population was lower than that of the B. thuringiensisserovar israelensis susceptible colony. We suggest that differences in protoxin proteolysis could contribute to the resistance of this Ae. aegypti colony.

Botulinum Toxin (Botox) for Facial Wrinkles

Science.gov (United States)

... Stories Español Eye Health / Eye Health A-Z Botulinum Toxin (Botox) for Facial Wrinkles Sections Botulinum Toxin (Botox) ... Facial Wrinkles How Does Botulinum Toxin (Botox) Work? Botulinum Toxin (Botox) for Facial Wrinkles Leer en Español: La ...

Generation and characterization of recombinant bivalent fusion protein r-Cpib for immunotherapy against Clostridium perfringens beta and iota toxemia.

Science.gov (United States)

Das, Shreya; Majumder, Saugata; Kingston, Joseph J; Batra, Harsh V

2016-02-01

Clostridium perfringens beta (CPB) and iota (CPI) toxaemias result in some of the most lethal forms of haemorrhagic and necrotic enteritis and sudden death syndrome affecting especially neonates. While CPB enterotoxemia is one of the most common forms of clostridial enterotoxemia, CPI enterotoxemia though putatively considered to be rare is an emerging cause of concern. The similarities in clinical manifestation, gross and histopathology findings of both types of toxaemias coupled to the infrequency of CPI toxaemia might lead to symptomatic misidentification with Type C resulting in therapeutic failure due to habitual administration of CPB anti-toxin which is ineffective against CPI. Therefore in the present study, to generate a composite anti-toxin capable of neutralizing both toxaemias, a novel bivalent chimera r-Cpib was constructed by splicing the non-toxic C terminal binding regions of CPB and CPI, via a flexible glycine linker (G4S) by overlap-extension PCR. The fusion protein was characterized for its therapeutic abilities toward CPI and CPB toxin neutralizations. The r-Cpib was found to be non-toxic and could competitively inhibit binding of CPB to host cell receptors thereby reducing its cytotoxicity. Immunization of mice with r-Cpib generated specific antibodies capable of neutralizing the above toxaemias both in vitro and in vivo. Caco-2 cells exposed to a mixture of anti-r-Cpib sera and native CPI or CPB, displayed significantly superior protection against the respective toxins while passive challenge of mice with a similar mixture resulted in 83 and 91% protection against CPI and CPB respectively. Alternatively, mice exposed to a mixture of sham sera and native toxins died within 2-3 days. This work thus demonstrates r-Cpib as a novel bivalent fusion protein capable of efficient immunotherapy against C. perfringens CPI and CPB toxaemia. Copyright © 2015 Elsevier Ltd. All rights reserved.

Engineering Venom’s Toxin-Neutralizing Antibody Fragments and Its Therapeutic Potential

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Larissa M. Alvarenga

2014-08-01

Full Text Available Serum therapy remains the only specific treatment against envenoming, but anti-venoms are still prepared by fragmentation of polyclonal antibodies isolated from hyper-immunized horse serum. Most of these anti-venoms are considered to be efficient, but their production is tedious, and their use may be associated with adverse effects. Recombinant antibodies and smaller functional units are now emerging as credible alternatives and constitute a source of still unexploited biomolecules capable of neutralizing venoms. This review will be a walk through the technologies that have recently been applied leading to novel antibody formats with better properties in terms of homogeneity, specific activity and possible safety.

Lymphocyte receptors for pertussis toxin

Energy Technology Data Exchange (ETDEWEB)

Clark, C.G.; Armstrong, G.D. (Univ. of Alberta, Edmonton (Canada))

1990-12-01

We have investigated human T-lymphocyte receptors for pertussis toxin by affinity isolation and photoaffinity labeling procedures. T lymphocytes were obtained from peripheral human blood, surface iodinated, and solubilized in Triton X-100. The iodinated mixture was then passed through pertussis toxin-agarose, and the fractions were analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Autoradiography of the fixed, dried gels revealed several bands in the pertussis toxin-bound fraction that were not observed in fractions obtained from histone or fetuin-agarose. Further investigations employed a photoaffinity labeling reagent, sulfosuccinimidyl 2-(p-azido-salicylamido)-1,3'-dithiopropionate, to identify pertussis toxin receptors in freshly isolated peripheral blood monocytic cells, T lymphocytes, and Jurkat cells. In all three cell systems, the pertussis toxin affinity probe specifically labeled a single protein species with an apparent molecular weight of 70,000 that was not observed when the procedure was performed in the presence of excess unmodified pertussis toxin. A protein comparable in molecular weight to the one detected by the photoaffinity labeling technique was also observed among the species that bound to pertussis toxin-agarose. The results suggest that pertussis toxin may bind to a 70,000-Da receptor in human T lymphocytes.

Topical botulinum toxin.

Science.gov (United States)

Collins, Ashley; Nasir, Adnan

2010-03-01

Nanotechnology is a rapidly growing discipline that capitalizes on the unique properties of matter engineered on the nanoscale. Vehicles incorporating nanotechnology have led to great strides in drug delivery, allowing for increased active ingredient stability, bioavailability, and site-specific targeting. Botulinum toxin has historically been used for the correction of neurological and neuromuscular disorders, such as torticollis, blepharospasm, and strabismus. Recent dermatological indications have been for the management of axillary hyperhydrosis and facial rhytides. Traditional methods of botulinum toxin delivery have been needle-based. These have been associated with increased pain and cost. Newer methods of botulinum toxin formulation have yielded topical preparations that are bioactive in small pilot clinical studies. While there are some risks associated with topical delivery, the refinement and standardization of delivery systems and techniques for the topical administration of botulinum toxin using nanotechnology is anticipated in the near future.

H5N1 whole-virus vaccine induces neutralizing antibodies in humans which are protective in a mouse passive transfer model.

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M Keith Howard

Full Text Available BACKGROUND: Vero cell culture-derived whole-virus H5N1 vaccines have been extensively tested in clinical trials and consistently demonstrated to be safe and immunogenic; however, clinical efficacy is difficult to evaluate in the absence of wide-spread human disease. A lethal mouse model has been utilized which allows investigation of the protective efficacy of active vaccination or passive transfer of vaccine induced sera following lethal H5N1 challenge. METHODS: We used passive transfer of immune sera to investigate antibody-mediated protection elicited by a Vero cell-derived, non-adjuvanted inactivated whole-virus H5N1 vaccine. Mice were injected intravenously with H5N1 vaccine-induced rodent or human immune sera and subsequently challenged with a lethal dose of wild-type H5N1 virus. RESULTS: Passive transfer of H5N1 vaccine-induced mouse, guinea pig and human immune sera provided dose-dependent protection of recipient mice against lethal challenge with wild-type H5N1 virus. Protective dose fifty values for serum H5N1 neutralizing antibody titers were calculated to be ≤1∶11 for all immune sera, independently of source species. CONCLUSIONS: These data underpin the confidence that the Vero cell culture-derived, whole-virus H5N1 vaccine will be effective in a pandemic situation and support the use of neutralizing serum antibody titers as a correlate of protection for H5N1 vaccines.

Non-lethal Clostridium sordellii bacteraemia in an immunocompromised patient with pleomorphic sarcoma.

Science.gov (United States)

Bonnecaze, Alex K; Stephens, Sarah Ellen Elza; Miller, Peter John

2016-08-03

Clostridium sordellii is a spore-forming anaerobic Gram-positive rod that has rarely been reported to cause disease in humans. Resultant mortality from infection is estimated at nearly 70% and is most often correlated with gynaecological procedures, intravenous drug abuse or trauma. C. sordellii infection often presents similarly to toxic shock syndrome (TSS); notable features of infection include refractory hypotension, haemoconcentration and marked leucocytosis. Although clinically similar to TSS, a notable difference is C. sordellii infections rarely involve fever. The organism's major toxins include haemorrhagic (TcsH) and lethal factor (TcsL), which function to disrupt cytoskeletal integrity. Current literature suggests treating C. sordelli infection with a broad-spectrum penicillin, metronidazole and clindamycin. We present a case of C. sordellii bacteraemia and septic shock in an immunocompromised patient who was recently diagnosed with pleomorphic gluteal sarcoma. Despite presenting in critical condition, the patient improved after aggressive hemodynamic resuscitation, source control and intravenous antibiotic therapy. 2016 BMJ Publishing Group Ltd.

Immuno-detection of cleaved SNAP-25 from differentiated mouse embryonic stem cells provides a sensitive assay for determination of botulinum A toxin and antitoxin potency.

Science.gov (United States)

Yadirgi, G; Stickings, P; Rajagopal, S; Liu, Y; Sesardic, D

2017-12-01

Botulinum toxin type A is a causative agent of human botulism. Due to high toxicity and ease of production it is classified by the Centres for Disease Control and Prevention as a category A bioterrorism agent. The same serotype, BoNT/A, is also the most widely used in pharmaceutical preparations for treatment of a diverse range of neuromuscular disorders. Traditionally, animals are used to confirm the presence and activity of toxin and to establish neutralizing capabilities of countermeasures in toxin neutralization tests. Cell based assays for BoNT/A have been reported as the most viable alternative to animal models, since they are capable of reflecting all key steps (binding, translocation, internalization and cleavage of intracellular substrate) involved in toxin activity. In this paper we report preliminary development of a simple immunochemical method for specifically detecting BoNT/A cleaved intracellular substrate, SNAP-25, in cell lysates of neurons derived from mouse embryonic stem cells. The assay offers sensitivity of better than 0.1LD50/ml (3fM) which is not matched by other functional assays, including the mouse bioassay, and provides serotype specificity for quantitative detection of BoNT/A and anti-BoNT/A antitoxin. Subject to formal validation, the method described here could potentially be used as a substitute for the mouse bioassay to measure potency and consistency of therapeutic products. Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.

Botulinum toxin injection - larynx

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Injection laryngoplasty; Botox - larynx: spasmodic dysphonia-BTX; Essential voice tremor (EVT)-btx; Glottic insufficiency; Percutaneous electromyography - guided botulinum toxin treatment; Percutaneous indirect laryngoscopy - guided botulinum toxin treatment; ...

Pre-clinical studies of toxin-specific Nanobodies: Evidence of in vivo efficacy to prevent fatal disturbances provoked by scorpion envenoming

International Nuclear Information System (INIS)

Hmila, Issam; Cosyns, Bernard; Tounsi, Hayfa; Roosens, Bram; Caveliers, Vicky; Abderrazek, Rahma Ben; Boubaker, Samir; Muyldermans, Serge; El Ayeb, Mohamed; Bouhaouala-Zahar, Balkiss; Lahoutte, Tony

2012-01-01

Scorpions represent a significant threat to humans and animals in various countries throughout the world. Recently, we introduced Nanobodies (Nbs) to combat more efficiently scorpion envenoming and demonstrated the performance of NbAahIF12 and NbAahII10 to neutralize scorpion toxins of Androctonus australis hector venom. A bispecific Nb construct (NbF12-10) comprising these two Nbs is far more protective than the classic Fab′ 2 based therapy and is the most efficient antivenom therapy against scorpion sting in preclinical studies. Now we investigate the biodistribution and pharmacokinetics of 99m Tc labeled Nbs by in vivo imaging in rodents and compared these data with those of the Fab′ 2 product (PAS). The pharmacodynamics of the Nbs was investigated in rats by in vivo echocardiography and it is shown that NbF12-10 prevents effectively the hemodynamic disturbances induced by a lethal dose of venom. Moreover, even a late injection of NbF12-10 restores the heart rate and brings the blood pressure to baseline values. Histology confirms that NbF12-10 prevents lung and heart lesions of treated mice after envenoming. In conjunction, in this preclinical study, we provide proof of concept that NbF12-10 prevents effectively the fatal disturbances induced by Androctonus venom, and that the Nanobody based therapeutic has a potential to substitute the classic Fab′ 2 based product as immunotherapeutic in scorpion envenoming. Further clinical study using larger cohorts of animals should be considered to confirm the full protecting potential of our NbF12-10. -- Highlights: ► Nanobody therapy prevents the hemodynamic disturbances induced by a lethal dose. ► Late injection of Nanobody restores hemodynamic parameters to baseline values. ► Nanobody therapy prevents lung and heart lesions of treated mice after envenoming. ► Labeled Nanobody and Fab’2 pharmacokinetics curves reach plateau in favour of Nanobody.

Pre-clinical studies of toxin-specific Nanobodies: Evidence of in vivo efficacy to prevent fatal disturbances provoked by scorpion envenoming

Energy Technology Data Exchange (ETDEWEB)

Hmila, Issam [Laboratoire des Venins et Toxines, Institut Pasteur de Tunis, 13 Place Pasteur, BP-74, 1002 Tunis (Tunisia); Cosyns, Bernard [Laboratory of In Vivo Cellular and Molecular Imaging, Vrije Universiteit Brussel (Belgium); Tounsi, Hayfa [Service d' Anatomo-Pathologie, Institut Pasteur de Tunis, 13 Place Pasteur, BP-74, 1002 Tunis (Tunisia); Roosens, Bram; Caveliers, Vicky [Laboratory of In Vivo Cellular and Molecular Imaging, Vrije Universiteit Brussel (Belgium); Abderrazek, Rahma Ben [Laboratoire des Venins et Toxines, Institut Pasteur de Tunis, 13 Place Pasteur, BP-74, 1002 Tunis (Tunisia); Boubaker, Samir [Service d' Anatomo-Pathologie, Institut Pasteur de Tunis, 13 Place Pasteur, BP-74, 1002 Tunis (Tunisia); Muyldermans, Serge [Laboratory of Cellular and Molecular Immunology, Vrije Universiteit Brussel, Pleinlaan 2, 1050 Brussel (Belgium); Department of Structural Biology, VIB, Brussels (Belgium); El Ayeb, Mohamed [Laboratoire des Venins et Toxines, Institut Pasteur de Tunis, 13 Place Pasteur, BP-74, 1002 Tunis (Tunisia); Bouhaouala-Zahar, Balkiss, E-mail: balkiss.bouhaouala@pasteur.rns.tn [Laboratoire des Venins et Toxines, Institut Pasteur de Tunis, 13 Place Pasteur, BP-74, 1002 Tunis (Tunisia); Faculté de Médecine de Tunis, Université de Tunis-El Manar (Tunisia); Lahoutte, Tony [Laboratory of In Vivo Cellular and Molecular Imaging, Vrije Universiteit Brussel (Belgium)

2012-10-15

Scorpions represent a significant threat to humans and animals in various countries throughout the world. Recently, we introduced Nanobodies (Nbs) to combat more efficiently scorpion envenoming and demonstrated the performance of NbAahIF12 and NbAahII10 to neutralize scorpion toxins of Androctonus australis hector venom. A bispecific Nb construct (NbF12-10) comprising these two Nbs is far more protective than the classic Fab′{sub 2} based therapy and is the most efficient antivenom therapy against scorpion sting in preclinical studies. Now we investigate the biodistribution and pharmacokinetics of {sup 99m}Tc labeled Nbs by in vivo imaging in rodents and compared these data with those of the Fab′{sub 2} product (PAS). The pharmacodynamics of the Nbs was investigated in rats by in vivo echocardiography and it is shown that NbF12-10 prevents effectively the hemodynamic disturbances induced by a lethal dose of venom. Moreover, even a late injection of NbF12-10 restores the heart rate and brings the blood pressure to baseline values. Histology confirms that NbF12-10 prevents lung and heart lesions of treated mice after envenoming. In conjunction, in this preclinical study, we provide proof of concept that NbF12-10 prevents effectively the fatal disturbances induced by Androctonus venom, and that the Nanobody based therapeutic has a potential to substitute the classic Fab′{sub 2} based product as immunotherapeutic in scorpion envenoming. Further clinical study using larger cohorts of animals should be considered to confirm the full protecting potential of our NbF12-10. -- Highlights: ► Nanobody therapy prevents the hemodynamic disturbances induced by a lethal dose. ► Late injection of Nanobody restores hemodynamic parameters to baseline values. ► Nanobody therapy prevents lung and heart lesions of treated mice after envenoming. ► Labeled Nanobody and Fab’2 pharmacokinetics curves reach plateau in favour of Nanobody.

Pyrethroids and Nectar Toxins Have Subtle Effects on the Motor Function, Grooming and Wing Fanning Behaviour of Honeybees (Apis mellifera).

Science.gov (United States)

Oliver, Caitlin J; Softley, Samantha; Williamson, Sally M; Stevenson, Philip C; Wright, Geraldine A

2015-01-01

Sodium channels, found ubiquitously in animal muscle cells and neurons, are one of the main target sites of many naturally-occurring, insecticidal plant compounds and agricultural pesticides. Pyrethroids, derived from compounds found only in the Asteraceae, are particularly toxic to insects and have been successfully used as pesticides including on flowering crops that are visited by pollinators. Pyrethrins, from which they were derived, occur naturally in the nectar of some flowering plant species. We know relatively little about how such compounds--i.e., compounds that target sodium channels--influence pollinators at low or sub-lethal doses. Here, we exposed individual adult forager honeybees to several compounds that bind to sodium channels to identify whether these compounds affect motor function. Using an assay previously developed to identify the effect of drugs and toxins on individual bees, we investigated how acute exposure to 10 ng doses (1 ppm) of the pyrethroid insecticides (cyfluthrin, tau-fluvalinate, allethrin and permethrin) and the nectar toxins (aconitine and grayanotoxin I) affected honeybee locomotion, grooming and wing fanning behaviour. Bees exposed to these compounds spent more time upside down and fanning their wings. They also had longer bouts of standing still. Bees exposed to the nectar toxin, aconitine, and the pyrethroid, allethrin, also spent less time grooming their antennae. We also found that the concentration of the nectar toxin, grayanotoxin I (GTX), fed to bees affected the time spent upside down (i.e., failure to perform the righting reflex). Our data show that low doses of pyrethroids and other nectar toxins that target sodium channels mainly influence motor function through their effect on the righting reflex of adult worker honeybees.

Mechanism of Diphtheria Toxin Catalytic Domain Delivery to the Eukaryotic Cell Cytosol and the Cellular Factors that Directly Participate in the Process

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Murphy, John R.

2011-01-01

Research on diphtheria and anthrax toxins over the past three decades has culminated in a detailed understanding of their structure function relationships (e.g., catalytic (C), transmembrane (T), and receptor binding (R) domains), as well as the identification of their eukaryotic cell surface receptor, an understanding of the molecular events leading to the receptor-mediated internalization of the toxin into an endosomal compartment, and the pH triggered conformational changes required for pore formation in the vesicle membrane. Recently, a major research effort has been focused on the development of a detailed understanding of the molecular interactions between each of these toxins and eukaryotic cell factors that play an essential role in the efficient translocation of their respective catalytic domains through the trans-endosomal vesicle membrane pore and delivery into the cell cytosol. In this review, I shall focus on recent findings that have led to a more detailed understanding of the mechanism by which the diphtheria toxin catalytic domain is delivered to the eukaryotic cell cytosol. While much work remains, it is becoming increasingly clear that the entry process is facilitated by specific interactions with a number of cellular factors in an ordered sequential fashion. In addition, since diphtheria, anthrax lethal factor and anthrax edema factor all carry multiple coatomer I complex binding motifs and COPI complex has been shown to play an essential role in entry process, it is likely that the initial steps in catalytic domain entry of these divergent toxins follow a common mechanism. PMID:22069710

Gender differences in onabotulinum toxin A dosing for adductor spasmodic dysphonia.

Science.gov (United States)

Lerner, Michael Z; Lerner, Benjamin A; Patel, Amit A; Blitzer, Andrew

2017-05-01

The objective of this study was to determine the influence of gender on onabotulinum toxin A dosing for the treatment of adductor spasmodic dysphonia symptoms. Retrospective review. A chart review of the senior author's database of botulinum toxin injections was performed. Patients diagnosed with adductor spasmodic dysphonia who received onabotulinum toxin A (BoNTA) injections to the thyroarytenoid muscle for at least 5 years were included for study. Patients who received alternate formulations of botulinum toxin (Myobloc, Dysport, or Xeomin) and patients with alternate diagnoses, such as abductor spasmodic dysphonia, tremor, and oromandibular dystonia, were excluded. The average BoNTA dose was calculated for each patient and statistical analysis was performed comparing the male and female groups. A total of 201 patients (52 males and 149 females) met inclusion criteria. The average follow-up times for the male and female groups were 10.2 ± 3.6 and 11.1 ± 4 years, respectively. The average BoNTA doses for the male and female groups were 0.6 ± 0.42 U and 1.3 ± 1.1 U, respectively. Statistical analysis was performed using an independent samples two-tailed t test yielding a P value of .0000000002. A large effect size was noted with Cohen's d = 0.85. The data from this retrospective chart review reveal a statistically and clinically significant correlation between female gender and higher average BoNTA dose for symptom control in adductor spasmodic dysphonia. Explanations for this observation are speculative and include a possible inverse relationship between optimal BoNTA dose and vocal fold mass and possibly greater neutralizing antibody formation among female patients. 4. Laryngoscope, 127:1131-1134, 2017. © 2016 The American Laryngological, Rhinological and Otological Society, Inc.

Efficacy of a potential trivalent vaccine based on Hc fragments of botulinum toxins A, B, and E produced in a cell-free expression system.

Science.gov (United States)

Zichel, R; Mimran, A; Keren, A; Barnea, A; Steinberger-Levy, I; Marcus, D; Turgeman, A; Reuveny, S

2010-05-01

Botulinum toxins produced by the anaerobic bacterium Clostridium botulinum are the most potent biological toxins in nature. Traditionally, people at risk are immunized with a formaldehyde-inactivated toxin complex. Second generation vaccines are based on the recombinant carboxy-terminal heavy-chain (Hc) fragment of the neurotoxin. However, the materialization of this approach is challenging, mainly due to the high AT content of clostridial genes. Herein, we present an alternative strategy in which the native genes encoding Hc proteins of botulinum toxins A, B, and E were used to express the recombinant Hc fragments in a cell-free expression system. We used the unique property of this open system to introduce different combinations of chaperone systems, protein disulfide isomerase (PDI), and reducing/oxidizing environments directly to the expression reaction. Optimized expression conditions led to increased production of soluble Hc protein, which was successfully scaled up using a continuous exchange (CE) cell-free system. Hc proteins were produced at a concentration of more than 1 mg/ml and purified by one-step Ni(+) affinity chromatography. Mice immunized with three injections containing 5 microg of any of the in vitro-expressed, alum-absorbed, Hc vaccines generated a serum enzyme-linked immunosorbent assay (ELISA) titer of 10(5) against the native toxin complex, which enabled protection against a high-dose toxin challenge (10(3) to 10(6) mouse 50% lethal dose [MsLD(50)]). Finally, immunization with a trivalent HcA, HcB, and HcE vaccine protected mice against the corresponding trivalent 10(5) MsLD(50) toxin challenge. Our results together with the latest developments in scalability of the in vitro protein expression systems offer alternative routes for the preparation of botulinum vaccine.

[Intoxication of botulinum toxin].

Science.gov (United States)

Chudzicka, Aleksandra

2015-09-01

Botulinum toxin is an egzotoxin produced by Gram positive bacteria Clostridium botulinum. It is among the most potent toxins known. The 3 main clinical presentations of botulism are as follows: foodborne botulism, infant botulism and wound botulism. The main symptom of intoxication is flat muscles paralysis. The treatment is supportive care and administration of antitoxin. In prevention the correct preparing of canned food is most important. Botulinum toxin is accepted as a biological weapon. © 2015 MEDPRESS.

Sensitivity of cancer cells to truncated diphtheria toxin.

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Yi Zhang

2010-05-01

Full Text Available Diphtheria toxin (DT has been utilized as a prospective anti-cancer agent for the targeted delivery of cytotoxic therapy to otherwise untreatable neoplasia. DT is an extremely potent toxin for which the entry of a single molecule into a cell can be lethal. DT has been targeted to cancer cells by deleting the cell receptor-binding domain and combining the remaining catalytic portion with targeting proteins that selectively bind to the surface of cancer cells. It has been assumed that "receptorless" DT cannot bind to and kill cells. In the present study, we report that "receptorless" recombinant DT385 is in fact cytotoxic to a variety of cancer cell lines.In vitro cytotoxicity of DT385 was measured by cell proliferation, cell staining and apoptosis assays. For in vivo studies, the chick chorioallantoic membrane (CAM system was used to evaluate the effect of DT385 on angiogenesis. The CAM and mouse model system was used to evaluate the effect of DT385 on HEp3 and Lewis lung carcinoma (LLC tumor growth, respectively.Of 18 human cancer cell lines tested, 15 were affected by DT385 with IC(50 ranging from 0.12-2.8 microM. Furthermore, high concentrations of DT385 failed to affect growth arrested cells. The cellular toxicity of DT385 was due to the inhibition of protein synthesis and induction of apoptosis. In vivo, DT385 diminished angiogenesis and decreased tumor growth in the CAM system, and inhibited the subcutaneous growth of LLC tumors in mice.DT385 possesses anti-angiogenic and anti-tumor activity and may have potential as a therapeutic agent.

Factors influencing circadian rhythms in acetaminophen lethality.

Science.gov (United States)

Schnell, R C; Bozigian, H P; Davies, M H; Merrick, B A; Park, K S; McMillan, D A

1984-01-01

Experiments were conducted to examine the effects of changes in lighting schedules and food consumption on circadian rhythms in acetaminophen lethality and hepatic glutathione levels in male mice. Under a normal lighting schedule (light: 06.00-18.00 h), male mice exhibited a circadian rhythm in acetaminophen lethality (peak: 18.00 h; nadir: 06.00, 10.00 h) and an inverse rhythm in hepatic glutathione concentrations (peak: 06.00, 10.00 h; nadir: 18.00 h). Under a reversed lighting schedule (light: 18.00-06.00 h) the glutathione rhythm was reversed and the rhythm in acetaminophen lethality was altered showing greater sensitivity to the drug. Under continuous light, there was a shift in the acetaminophen lethality and the hepatic glutathione rhythms. Under continuous dark, both rhythms were abolished. Under a normal lighting regimen, hepatic glutathione levels were closely correlated with food consumption; i.e., both were increased during the dark phase and decreased during the light phase. Fasting the mice for 12 h abolished the rhythms in acetaminophen lethality and hepatic glutathione levels; moreover, the lethality was increased and the hepatic glutathione levels were decreased. These experiments show that both lighting schedules and feeding can alter the circadian rhythms in acetaminophen lethality and hepatic glutathione levels in male mice.

Low sensitivity of fecal toxin A/B enzyme immunoassay for diagnosis of Clostridium difficile infection in immunocompromised patients.

Science.gov (United States)

Erb, S; Frei, R; Strandén, A M; Dangel, M; Tschudin-Sutter, S; Widmer, A F

2015-11-01

The optimal approach in laboratory diagnosis of Clostridium difficile infection (CDI) is still not well defined. Toxigenic culture (TC) or alternatively fecal toxin assay by cell cytotoxicity neutralization assay are considered to be the reference standard, but these methods are time-consuming and labor intensive. In many medical centers, diagnosis of CDI is therefore still based on fecal toxin A/B enzyme immunoassay (EIA) directly from stool alone, balancing cost and speed against limited diagnostic sensitivity. The aim of the study was to assess in which patient population the additional workload of TC is justified. All consecutive stool specimens submitted for diagnosis of suspected CDI between 2004 and 2011 at a tertiary-care center were examined by toxin EIA and TC. Clinical data of patients with established diagnosis of CDI were collected in a standardized case-report form. From 12,481 stool specimens submitted to the microbiologic laboratory, 480 (3.8%) fulfilled CDI criteria; 274 (57.1%) were diagnosed by toxin EIA; and an additional 206 (42.9%) were diagnosed by TC when toxin EIA was negative. Independent predictors for negative toxin EIA but positive TC were high-dose corticosteroids (odds ratio (OR) 2.97, 95% confidence interval (CI) 1.50-5.90, p 0.002), leukocytopenia <1000/μL (OR 2.52, 95% CI 1.22-5.23, p 0.013) and nonsevere CDI (OR 2.21, 95% CI 1.39-3.50, p 0.001). There was no difference in outcomes such as in-hospital mortality and recurrence between both groups. In conclusion, negative toxin EIA does not rule out CDI in immunocompromised patients in the setting of relevant clinical symptoms. Methods with improved sensitivity such as TC or PCR should be used, particularly in this patient population. Copyright © 2015 European Society of Clinical Microbiology and Infectious Diseases. Published by Elsevier Ltd. All rights reserved.

Plant Insecticidal Toxins in Ecological Networks

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Sébastien Ibanez

2012-04-01

Full Text Available Plant secondary metabolites play a key role in plant-insect interactions, whether constitutive or induced, C- or N-based. Anti-herbivore defences against insects can act as repellents, deterrents, growth inhibitors or cause direct mortality. In turn, insects have evolved a variety of strategies to act against plant toxins, e.g., avoidance, excretion, sequestration and degradation of the toxin, eventually leading to a co-evolutionary arms race between insects and plants and to co-diversification. Anti-herbivore defences also negatively impact mutualistic partners, possibly leading to an ecological cost of toxin production. However, in other cases toxins can also be used by plants involved in mutualistic interactions to exclude inadequate partners and to modify the cost/benefit ratio of mutualism to their advantage. When considering the whole community, toxins have an effect at many trophic levels. Aposematic insects sequester toxins to defend themselves against predators. Depending on the ecological context, toxins can either increase insects’ vulnerability to parasitoids and entomopathogens or protect them, eventually leading to self-medication. We conclude that studying the community-level impacts of plant toxins can provide new insights into the synthesis between community and evolutionary ecology.

Plant insecticidal toxins in ecological networks.

Science.gov (United States)

Ibanez, Sébastien; Gallet, Christiane; Després, Laurence

2012-04-01

Plant secondary metabolites play a key role in plant-insect interactions, whether constitutive or induced, C- or N-based. Anti-herbivore defences against insects can act as repellents, deterrents, growth inhibitors or cause direct mortality. In turn, insects have evolved a variety of strategies to act against plant toxins, e.g., avoidance, excretion, sequestration and degradation of the toxin, eventually leading to a co-evolutionary arms race between insects and plants and to co-diversification. Anti-herbivore defences also negatively impact mutualistic partners, possibly leading to an ecological cost of toxin production. However, in other cases toxins can also be used by plants involved in mutualistic interactions to exclude inadequate partners and to modify the cost/benefit ratio of mutualism to their advantage. When considering the whole community, toxins have an effect at many trophic levels. Aposematic insects sequester toxins to defend themselves against predators. Depending on the ecological context, toxins can either increase insects' vulnerability to parasitoids and entomopathogens or protect them, eventually leading to self-medication. We conclude that studying the community-level impacts of plant toxins can provide new insights into the synthesis between community and evolutionary ecology.

Polymorphic toxin systems: Comprehensive characterization of trafficking modes, processing, mechanisms of action, immunity and ecology using comparative genomics

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Zhang Dapeng

2012-06-01

Full Text Available Abstract Background Proteinaceous toxins are observed across all levels of inter-organismal and intra-genomic conflicts. These include recently discovered prokaryotic polymorphic toxin systems implicated in intra-specific conflicts. They are characterized by a remarkable diversity of C-terminal toxin domains generated by recombination with standalone toxin-coding cassettes. Prior analysis revealed a striking diversity of nuclease and deaminase domains among the toxin modules. We systematically investigated polymorphic toxin systems using comparative genomics, sequence and structure analysis. Results Polymorphic toxin systems are distributed across all major bacterial lineages and are delivered by at least eight distinct secretory systems. In addition to type-II, these include type-V, VI, VII (ESX, and the poorly characterized “Photorhabdus virulence cassettes (PVC”, PrsW-dependent and MuF phage-capsid-like systems. We present evidence that trafficking of these toxins is often accompanied by autoproteolytic processing catalyzed by HINT, ZU5, PrsW, caspase-like, papain-like, and a novel metallopeptidase associated with the PVC system. We identified over 150 distinct toxin domains in these systems. These span an extraordinary catalytic spectrum to include 23 distinct clades of peptidases, numerous previously unrecognized versions of nucleases and deaminases, ADP-ribosyltransferases, ADP ribosyl cyclases, RelA/SpoT-like nucleotidyltransferases, glycosyltranferases and other enzymes predicted to modify lipids and carbohydrates, and a pore-forming toxin domain. Several of these toxin domains are shared with host-directed effectors of pathogenic bacteria. Over 90 families of immunity proteins might neutralize anywhere between a single to at least 27 distinct types of toxin domains. In some organisms multiple tandem immunity genes or immunity protein domains are organized into polyimmunity loci or polyimmunity proteins. Gene-neighborhood-analysis of

Exploration of immunoglobulin transcriptomes from mice immunized with three-finger toxins and phospholipases A2 from the Central American coral snake, Micrurus nigrocinctus

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Andreas H. Laustsen

2017-01-01

Full Text Available Snakebite envenomings represent a neglected public health issue in many parts of the rural tropical world. Animal-derived antivenoms have existed for more than a hundred years and are effective in neutralizing snake venom toxins when timely administered. However, the low immunogenicity of many small but potent snake venom toxins represents a challenge for obtaining a balanced immune response against the medically relevant components of the venom. Here, we employ high-throughput sequencing of the immunoglobulin (Ig transcriptome of mice immunized with a three-finger toxin and a phospholipase A2 from the venom of the Central American coral snake, Micrurus nigrocinctus. Although exploratory in nature, our indicate results showed that only low frequencies of mRNA encoding IgG isotypes, the most relevant isotype for therapeutic purposes, were present in splenocytes of five mice immunized with 6 doses of the two types of toxins over 90 days. Furthermore, analysis of Ig heavy chain transcripts showed that no particular combination of variable (V and joining (J gene segments had been selected in the immunization process, as would be expected after a strong humoral immune response to a single antigen. Combined with the titration of toxin-specific antibodies in the sera of immunized mice, these data support the low immunogenicity of three-finger toxins and phospholipases A2found in M. nigrocinctusvenoms, and highlight the need for future studies analyzing the complexity of antibody responses to toxins at the molecular level.

Lethal mutants and truncated selection together solve a paradox of the origin of life.

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David B Saakian

Full Text Available BACKGROUND: Many attempts have been made to describe the origin of life, one of which is Eigen's cycle of autocatalytic reactions [Eigen M (1971 Naturwissenschaften 58, 465-523], in which primordial life molecules are replicated with limited accuracy through autocatalytic reactions. For successful evolution, the information carrier (either RNA or DNA or their precursor must be transmitted to the next generation with a minimal number of misprints. In Eigen's theory, the maximum chain length that could be maintained is restricted to 100-1000 nucleotides, while for the most primitive genome the length is around 7000-20,000. This is the famous error catastrophe paradox. How to solve this puzzle is an interesting and important problem in the theory of the origin of life. METHODOLOGY/PRINCIPAL FINDINGS: We use methods of statistical physics to solve this paradox by carefully analyzing the implications of neutral and lethal mutants, and truncated selection (i.e., when fitness is zero after a certain Hamming distance from the master sequence for the critical chain length. While neutral mutants play an important role in evolution, they do not provide a solution to the paradox. We have found that lethal mutants and truncated selection together can solve the error catastrophe paradox. There is a principal difference between prebiotic molecule self-replication and proto-cell self-replication stages in the origin of life. CONCLUSIONS/SIGNIFICANCE: We have applied methods of statistical physics to make an important breakthrough in the molecular theory of the origin of life. Our results will inspire further studies on the molecular theory of the origin of life and biological evolution.

Synthesis and biology of cyclic imine toxins, an emerging class of potent, globally distributed marine toxins.

Science.gov (United States)

Stivala, Craig E; Benoit, Evelyne; Aráoz, Rómulo; Servent, Denis; Novikov, Alexei; Molgó, Jordi; Zakarian, Armen

2015-03-01

From a small group of exotic compounds isolated only two decades ago, Cyclic Imine (CI) toxins have become a major class of marine toxins with global distribution. Their distinct chemical structure, biological mechanism of action, and intricate chemistry ensures that CI toxins will continue to be the subject of fascinating fundamental studies in the broad fields of chemistry, chemical biology, and toxicology. The worldwide occurrence of potent CI toxins in marine environments, their accumulation in shellfish, and chemical stability are important considerations in assessing risk factors for human health. This review article aims to provide an account of chemistry, biology, and toxicology of CI toxins from their discovery to the present day.

Defense against Toxin Weapons

National Research Council Canada - National Science Library

Franz, David

1998-01-01

.... We typically fear what we do not understand. Although un- derstanding toxin poisoning is less useful in a toxin attack than knowledge of cold injury on an Arctic battlefield, information on any threat reduces its potential to harm...

Food toxin detection with atomic force microscope

Science.gov (United States)

Externally introduced toxins or internal spoilage correlated pathogens and their metabolites are all potential sources of food toxins. To prevent and protect unsafe food, many food toxin detection techniques have been developed to detect various toxins for quality control. Although several routine m...

Cellular Entry of Clostridium perfringens Iota-Toxin and Clostridium botulinum C2 Toxin.

Science.gov (United States)

Takehara, Masaya; Takagishi, Teruhisa; Seike, Soshi; Oda, Masataka; Sakaguchi, Yoshihiko; Hisatsune, Junzo; Ochi, Sadayuki; Kobayashi, Keiko; Nagahama, Masahiro

2017-08-11

Clostridium perfringens iota-toxin and Clostridium botulinum C2 toxin are composed of two non-linked proteins, one being the enzymatic component and the other being the binding/translocation component. These latter components recognize specific receptors and oligomerize in plasma membrane lipid-rafts, mediating the uptake of the enzymatic component into the cytosol. Enzymatic components induce actin cytoskeleton disorganization through the ADP-ribosylation of actin and are responsible for cell rounding and death. This review focuses upon the recent advances in cellular internalization of clostridial binary toxins.

Conditional Toxin Splicing Using a Split Intein System.

Science.gov (United States)

Alford, Spencer C; O'Sullivan, Connor; Howard, Perry L

2017-01-01

Protein toxin splicing mediated by split inteins can be used as a strategy for conditional cell ablation. The approach requires artificial fragmentation of a potent protein toxin and tethering each toxin fragment to a split intein fragment. The toxin-intein fragments are, in turn, fused to dimerization domains, such that addition of a dimerizing agent reconstitutes the split intein. These chimeric toxin-intein fusions remain nontoxic until the dimerizer is added, resulting in activation of intein splicing and ligation of toxin fragments to form an active toxin. Considerations for the engineering and implementation of conditional toxin splicing (CTS) systems include: choice of toxin split site, split site (extein) chemistry, and temperature sensitivity. The following method outlines design criteria and implementation notes for CTS using a previously engineered system for splicing a toxin called sarcin, as well as for developing alternative CTS systems.

Mutant with diphtheria toxin receptor and acidification function but defective in entry of toxin

International Nuclear Information System (INIS)

Kohno, Kenji; Hayes, H.; Mekada, Eisuke; Uchida, Tsuyoshi

1987-01-01

A mutant of Chinese hamster ovary cells, GE1, that is highly resistant to diphtheria toxin was isolated. The mutant contains 50% ADP-ribosylatable elongation factor 2, but its protein synthesis was not inhibited by the toxin even at concentrations above 100 μg/ml. 125 I-labeled diphtheria toxin was associated with GE1 cells as well as with the parent cells but did not block protein synthesis of GE1 cells even when the cells were exposed to low pH in the presence or absence of NH 4 Cl. The infections of GE1 cells and the parent cells by vesicular stomatitis virus were similar. GE1 cells were cross-resistant to Pseudomonas aeruginosa exotoxin A and so were about 1,000 times more resistant to this toxin than the parent cells. Hybrids of GE1 cells and the parent cells or mutant cells lacking a functional receptor were more sensitive to diphtheria toxin than GE1 cells. These results suggest that entry of diphtheria toxin into cells requires a cellular factor(s) in addition to those involved in receptor function and acidification of endosomes and that GE1 cells do not express this cellular factor. This character is recessive in GE1 cells

Infectious Entry and Neutralization of Pathogenic JC Polyomaviruses

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Eileen M. Geoghegan

2017-10-01

Full Text Available Summary: Progressive multifocal leukoencephalopathy (PML is a lethal brain disease caused by uncontrolled replication of JC polyomavirus (JCV. JCV strains recovered from the brains of PML patients carry mutations that prevent the engagement of sialylated glycans, which are thought to serve as receptors for the infectious entry of wild-type JCV. In this report, we show that non-sialylated glycosaminoglycans (GAGs can serve as alternative attachment receptors for the infectious entry of both wild-type and PML mutant JCV strains. After GAG-mediated attachment, PML mutant strains engage non-sialylated non-GAG co-receptor glycans, such as asialo-GM1. JCV-neutralizing monoclonal antibodies isolated from patients who recovered from PML appear to block infection by preventing the docking of post-attachment co-receptor glycans in an apical pocket of the JCV major capsid protein. Identification of the GAG-dependent/sialylated glycan-independent alternative entry pathway should facilitate the development of infection inhibitors, including recombinant neutralizing antibodies. : Geoghegan et al. show that JC polyomavirus strains that cause brain disease infect cells via a pathway involving a heparin-like attachment receptor and a non-sialylated co-receptor. Candidate therapeutic human monoclonal antibodies neutralize by blocking co-receptor engagement. Keywords: polyomavirus, JC, BK, SV40, progressive multifocal leukoencephalopathy, PML, monoclonal antibody, mAb, virus entry, receptor

Monalysin, a novel ß-pore-forming toxin from the Drosophila pathogen Pseudomonas entomophila, contributes to host intestinal damage and lethality.

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Onya Opota

2011-09-01

Full Text Available Pseudomonas entomophila is an entomopathogenic bacterium that infects and kills Drosophila. P. entomophila pathogenicity is linked to its ability to cause irreversible damages to the Drosophila gut, preventing epithelium renewal and repair. Here we report the identification of a novel pore-forming toxin (PFT, Monalysin, which contributes to the virulence of P. entomophila against Drosophila. Our data show that Monalysin requires N-terminal cleavage to become fully active, forms oligomers in vitro, and induces pore-formation in artificial lipid membranes. The prediction of the secondary structure of the membrane-spanning domain indicates that Monalysin is a PFT of the ß-type. The expression of Monalysin is regulated by both the GacS/GacA two-component system and the Pvf regulator, two signaling systems that control P. entomophila pathogenicity. In addition, AprA, a metallo-protease secreted by P. entomophila, can induce the rapid cleavage of pro-Monalysin into its active form. Reduced cell death is observed upon infection with a mutant deficient in Monalysin production showing that Monalysin plays a role in P. entomophila ability to induce intestinal cell damages, which is consistent with its activity as a PFT. Our study together with the well-established action of Bacillus thuringiensis Cry toxins suggests that production of PFTs is a common strategy of entomopathogens to disrupt insect gut homeostasis.

Computational Studies of Snake Venom Toxins.

Science.gov (United States)

Ojeda, Paola G; Ramírez, David; Alzate-Morales, Jans; Caballero, Julio; Kaas, Quentin; González, Wendy

2017-12-22

Most snake venom toxins are proteins, and participate to envenomation through a diverse array of bioactivities, such as bleeding, inflammation, and pain, cytotoxic, cardiotoxic or neurotoxic effects. The venom of a single snake species contains hundreds of toxins, and the venoms of the 725 species of venomous snakes represent a large pool of potentially bioactive proteins. Despite considerable discovery efforts, most of the snake venom toxins are still uncharacterized. Modern bioinformatics tools have been recently developed to mine snake venoms, helping focus experimental research on the most potentially interesting toxins. Some computational techniques predict toxin molecular targets, and the binding mode to these targets. This review gives an overview of current knowledge on the ~2200 sequences, and more than 400 three-dimensional structures of snake toxins deposited in public repositories, as well as of molecular modeling studies of the interaction between these toxins and their molecular targets. We also describe how modern bioinformatics have been used to study the snake venom protein phospholipase A2, the small basic myotoxin Crotamine, and the three-finger peptide Mambalgin.

Computational Studies of Snake Venom Toxins

Directory of Open Access Journals (Sweden)

Paola G. Ojeda

2017-12-01

Full Text Available Most snake venom toxins are proteins, and participate to envenomation through a diverse array of bioactivities, such as bleeding, inflammation, and pain, cytotoxic, cardiotoxic or neurotoxic effects. The venom of a single snake species contains hundreds of toxins, and the venoms of the 725 species of venomous snakes represent a large pool of potentially bioactive proteins. Despite considerable discovery efforts, most of the snake venom toxins are still uncharacterized. Modern bioinformatics tools have been recently developed to mine snake venoms, helping focus experimental research on the most potentially interesting toxins. Some computational techniques predict toxin molecular targets, and the binding mode to these targets. This review gives an overview of current knowledge on the ~2200 sequences, and more than 400 three-dimensional structures of snake toxins deposited in public repositories, as well as of molecular modeling studies of the interaction between these toxins and their molecular targets. We also describe how modern bioinformatics have been used to study the snake venom protein phospholipase A2, the small basic myotoxin Crotamine, and the three-finger peptide Mambalgin.

Cellular Entry of Clostridium perfringens Iota-Toxin and Clostridium botulinum C2 Toxin

Directory of Open Access Journals (Sweden)

Masaya Takehara

2017-08-01

Full Text Available Clostridium perfringens iota-toxin and Clostridium botulinum C2 toxin are composed of two non-linked proteins, one being the enzymatic component and the other being the binding/translocation component. These latter components recognize specific receptors and oligomerize in plasma membrane lipid-rafts, mediating the uptake of the enzymatic component into the cytosol. Enzymatic components induce actin cytoskeleton disorganization through the ADP-ribosylation of actin and are responsible for cell rounding and death. This review focuses upon the recent advances in cellular internalization of clostridial binary toxins.

EGA Protects Mammalian Cells from Clostridium difficile CDT, Clostridium perfringens Iota Toxin and Clostridium botulinum C2 Toxin.

Science.gov (United States)

Schnell, Leonie; Mittler, Ann-Katrin; Sadi, Mirko; Popoff, Michel R; Schwan, Carsten; Aktories, Klaus; Mattarei, Andrea; Azarnia Tehran, Domenico; Montecucco, Cesare; Barth, Holger

2016-04-01

The pathogenic bacteria Clostridium difficile, Clostridium perfringens and Clostridium botulinum produce the binary actin ADP-ribosylating toxins CDT, iota and C2, respectively. These toxins are composed of a transport component (B) and a separate enzyme component (A). When both components assemble on the surface of mammalian target cells, the B components mediate the entry of the A components via endosomes into the cytosol. Here, the A components ADP-ribosylate G-actin, resulting in depolymerization of F-actin, cell-rounding and eventually death. In the present study, we demonstrate that 4-bromobenzaldehyde N-(2,6-dimethylphenyl)semicarbazone (EGA), a compound that protects cells from multiple toxins and viruses, also protects different mammalian epithelial cells from all three binary actin ADP-ribosylating toxins. In contrast, EGA did not inhibit the intoxication of cells with Clostridium difficile toxins A and B, indicating a possible different entry route for this toxin. EGA does not affect either the binding of the C2 toxin to the cells surface or the enzyme activity of the A components of CDT, iota and C2, suggesting that this compound interferes with cellular uptake of the toxins. Moreover, for C2 toxin, we demonstrated that EGA inhibits the pH-dependent transport of the A component across cell membranes. EGA is not cytotoxic, and therefore, we propose it as a lead compound for the development of novel pharmacological inhibitors against clostridial binary actin ADP-ribosylating toxins.

EGA Protects Mammalian Cells from Clostridium difficile CDT, Clostridium perfringens Iota Toxin and Clostridium botulinum C2 Toxin

Science.gov (United States)

Schnell, Leonie; Mittler, Ann-Katrin; Sadi, Mirko; Popoff, Michel R.; Schwan, Carsten; Aktories, Klaus; Mattarei, Andrea; Tehran, Domenico Azarnia; Montecucco, Cesare; Barth, Holger

2016-01-01

The pathogenic bacteria Clostridium difficile, Clostridium perfringens and Clostridium botulinum produce the binary actin ADP-ribosylating toxins CDT, iota and C2, respectively. These toxins are composed of a transport component (B) and a separate enzyme component (A). When both components assemble on the surface of mammalian target cells, the B components mediate the entry of the A components via endosomes into the cytosol. Here, the A components ADP-ribosylate G-actin, resulting in depolymerization of F-actin, cell-rounding and eventually death. In the present study, we demonstrate that 4-bromobenzaldehyde N-(2,6-dimethylphenyl)semicarbazone (EGA), a compound that protects cells from multiple toxins and viruses, also protects different mammalian epithelial cells from all three binary actin ADP-ribosylating toxins. In contrast, EGA did not inhibit the intoxication of cells with Clostridium difficile toxins A and B, indicating a possible different entry route for this toxin. EGA does not affect either the binding of the C2 toxin to the cells surface or the enzyme activity of the A components of CDT, iota and C2, suggesting that this compound interferes with cellular uptake of the toxins. Moreover, for C2 toxin, we demonstrated that EGA inhibits the pH-dependent transport of the A component across cell membranes. EGA is not cytotoxic, and therefore, we propose it as a lead compound for the development of novel pharmacological inhibitors against clostridial binary actin ADP-ribosylating toxins. PMID:27043629

Deadly hairs, lethal feathers--convergent evolution of poisonous integument in mammals and birds.

Science.gov (United States)

Plikus, Maksim V; Astrowski, Aliaksandr A

2014-07-01

Hairs and feathers are textbook examples of the convergent evolution of the follicular appendage structure between mammals and birds. While broadly recognized for their convergent thermoregulatory, camouflage and sexual display functions, hairs and feathers are rarely thought of as deadly defence tools. Several recent studies, however, show that in some species of mammals and birds, the integument can, in fact, be a de facto lethal weapon. One mammalian example is provided by African crested rats, which seek for and chew on the bark of plants containing the highly potent toxin, ouabain. These rats then coat their fur with ouabain-containing saliva. For efficient toxin retention, the rodents have evolved highly specialized fenestrated and mostly hollow hair shafts that soak up liquids, which essentially function as wicks. On the avian side of the vertebrate integumental variety spectrum, several species of birds of New Guinea have evolved resistance to highly potent batrachotoxins, which they acquire from their insect diet. While the mechanism of bird toxicity remains obscure, in a recently published issue of the journal, Dumbacher and Menon explore the intriguing idea that to achieve efficient storage of batrachotoxins in their skin, some birds exploit the basic permeability barrier function of their epidermis. Batrachotoxins become preferentially sequestered in their epidermis and are then transferred to feathers, likely through the exploitation of specialized avian lipid-storing multigranular body organelles. Here, we discuss wider implications of this intriguing concept. © 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Lethal and sub-lethal effects of five pesticides used in rice farming on the earthworm Eisenia fetida

NARCIS (Netherlands)

Rico, Andreu; Sabater, Consuelo; Castillo, María Ángeles

2016-01-01

The toxicity of five pesticides typically used in rice farming (trichlorfon, dimethoate, carbendazim, tebuconazole and prochloraz) was evaluated on different lethal and sub-lethal endpoints of the earthworm Eisenia fetida. The evaluated endpoints included: avoidance behaviour after an exposure

Lethal Epistaxis.

Science.gov (United States)

Byard, Roger W

2016-09-01

Epistaxis or nosebleed refers to bleeding from the nostrils, nasal cavity, or nasopharynx. Occasional cases may present with torrential lethal hemorrhage. Three cases are reported to demonstrate particular features: Case 1: A 51-year-old woman with lethal epistaxis with no obvious bleeding source; Case 2: A 77-year-old man with treated nasopharyngeal carcinoma who died from epistaxis arising from a markedly neovascularized tumor bed; Case 3: A 2-year-old boy with hemophilia B who died from epistaxis with airway obstruction in addition to gastrointestinal bleeding. Epistaxis may be associated with trauma, tumors, vascular malformations, bleeding diatheses, infections, pregnancy, endometriosis, and a variety of different drugs. Careful dissection of the nasal cavity is required to locate the site of hemorrhage and to identify any predisposing conditions. This may be guided by postmortem computerized tomographic angiography (PCTA). Despite careful dissection, however, a source of bleeding may never be identified. © 2016 American Academy of Forensic Sciences.

SVM-based prediction of propeptide cleavage sites in spider toxins identifies toxin innovation in an Australian tarantula.

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Emily S W Wong

Full Text Available Spider neurotoxins are commonly used as pharmacological tools and are a popular source of novel compounds with therapeutic and agrochemical potential. Since venom peptides are inherently toxic, the host spider must employ strategies to avoid adverse effects prior to venom use. It is partly for this reason that most spider toxins encode a protective proregion that upon enzymatic cleavage is excised from the mature peptide. In order to identify the mature toxin sequence directly from toxin transcripts, without resorting to protein sequencing, the propeptide cleavage site in the toxin precursor must be predicted bioinformatically. We evaluated different machine learning strategies (support vector machines, hidden Markov model and decision tree and developed an algorithm (SpiderP for prediction of propeptide cleavage sites in spider toxins. Our strategy uses a support vector machine (SVM framework that combines both local and global sequence information. Our method is superior or comparable to current tools for prediction of propeptide sequences in spider toxins. Evaluation of the SVM method on an independent test set of known toxin sequences yielded 96% sensitivity and 100% specificity. Furthermore, we sequenced five novel peptides (not used to train the final predictor from the venom of the Australian tarantula Selenotypus plumipes to test the accuracy of the predictor and found 80% sensitivity and 99.6% 8-mer specificity. Finally, we used the predictor together with homology information to predict and characterize seven groups of novel toxins from the deeply sequenced venom gland transcriptome of S. plumipes, which revealed structural complexity and innovations in the evolution of the toxins. The precursor prediction tool (SpiderP is freely available on ArachnoServer (http://www.arachnoserver.org/spiderP.html, a web portal to a comprehensive relational database of spider toxins. All training data, test data, and scripts used are available from

CD4 T cell-mediated protection from lethal influenza: perforin and antibody-mediated mechanisms give a one-two punch.

Science.gov (United States)

Brown, Deborah M; Dilzer, Allison M; Meents, Dana L; Swain, Susan L

2006-09-01

The mechanisms whereby CD4 T cells contribute to the protective response against lethal influenza infection remain poorly characterized. To define the role of CD4 cells in protection against a highly pathogenic strain of influenza, virus-specific TCR transgenic CD4 effectors were generated in vitro and transferred into mice given lethal influenza infection. Primed CD4 effectors conferred protection against lethal infection over a broad range of viral dose. The protection mediated by CD4 effectors did not require IFN-gamma or host T cells, but did result in increased anti-influenza Ab titers compared with untreated controls. Further studies indicated that CD4-mediated protection at high doses of influenza required B cells, and that passive transfer of anti-influenza immune serum was therapeutic in B cell-deficient mice, but only when CD4 effectors were present. Primed CD4 cells also acquired perforin (Pfn)-mediated cytolytic activity during effector generation, suggesting a second mechanism used by CD4 cells to confer protection. Pfn-deficient CD4 effectors were less able to promote survival in intact BALB/c mice and were unable to provide protection in B cell-deficient mice, indicating that Ab-independent protection by CD4 effectors requires Pfn. Therefore, CD4 effectors mediate protection to lethal influenza through at least two mechanisms: Pfn-mediated cytotoxicity early in the response promoted survival independently of Ab production, whereas CD4-driven B cell responses resulted in high titer Abs that neutralized remaining virus.

The Effector Domain Region of the Vibrio vulnificus MARTX Toxin Confers Biphasic Epithelial Barrier Disruption and Is Essential for Systemic Spread from the Intestine.

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Hannah E Gavin

2017-01-01

Full Text Available Vibrio vulnificus causes highly lethal bacterial infections in which the Multifunctional Autoprocessing Repeats-in-Toxins (MARTX toxin product of the rtxA1 gene is a key virulence factor. MARTX toxins are secreted proteins up to 5208 amino acids in size. Conserved MARTX N- and C-terminal repeat regions work in concert to form pores in eukaryotic cell membranes, through which the toxin's central region of modular effector domains is translocated. Upon inositol hexakisphosphate-induced activation of the of the MARTX cysteine protease domain (CPD in the eukaryotic cytosol, effector domains are released from the holotoxin by autoproteolytic activity. We previously reported that the native MARTX toxin effector domain repertoire is dispensable for epithelial cellular necrosis in vitro, but essential for cell rounding and apoptosis prior to necrotic cell death. Here we use an intragastric mouse model to demonstrate that the effector domain region is required for bacterial virulence during intragastric infection. The MARTX effector domain region is essential for bacterial dissemination from the intestine, but dissemination occurs in the absence of overt intestinal tissue pathology. We employ an in vitro model of V. vulnificus interaction with polarized colonic epithelial cells to show that the MARTX effector domain region induces rapid intestinal barrier dysfunction and increased paracellular permeability prior to onset of cell lysis. Together, these results negate the inherent assumption that observations of necrosis in vitro directly predict bacterial virulence, and indicate a paradigm shift in our conceptual understanding of MARTX toxin function during intestinal infection. Results implicate the MARTX effector domain region in mediating early bacterial dissemination from the intestine to distal organs-a key step in V. vulnificus foodborne pathogenesis-even before onset of overt intestinal pathology.

Microalgal toxin(s): characteristics and importance

African Journals Online (AJOL)

Prokaryotic and eukaryotic microalgae produce a wide array of compounds with biological activities. These include antibiotics, algicides, toxins, pharmaceutically active compounds and plant growth regulators. Toxic microalgae, in this sense, are common only among the cyanobacteria and dinoflagellates. The microalgal ...

Immunotoxins: The Role of the Toxin

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David FitzGerald

2013-08-01

Full Text Available Immunotoxins are antibody-toxin bifunctional molecules that rely on intracellular toxin action to kill target cells. Target specificity is determined via the binding attributes of the chosen antibody. Mostly, but not exclusively, immunotoxins are purpose-built to kill cancer cells as part of novel treatment approaches. Other applications for immunotoxins include immune regulation and the treatment of viral or parasitic diseases. Here we discuss the utility of protein toxins, of both bacterial and plant origin, joined to antibodies for targeting cancer cells. Finally, while clinical goals are focused on the development of novel cancer treatments, much has been learned about toxin action and intracellular pathways. Thus toxins are considered both medicines for treating human disease and probes of cellular function.

Lethal congenital contracture syndrome (LCCS) and other lethal arthrogryposes in Finland--an epidemiological study.

Science.gov (United States)

Pakkasjärvi, Niklas; Ritvanen, Annukka; Herva, Riitta; Peltonen, Leena; Kestilä, Marjo; Ignatius, Jaakko

2006-09-01

Arthrogryposis multiplex congenita is a heterogeneous group of disorders characterized by multiple contractures with an estimated frequency of 1 in 3,000 births. With improving diagnostic methods, increasing numbers of fetuses with arthrogryposis are found. The pathogenetic mechanisms are relatively well known but the epidemiology and genetics of the prenatally lethal forms of arthrogryposis are less well known. In this study we collected all cases of a multiple contractures diagnosed in Finland during 1987-2002 including live born infants, stillbirths, and terminated pregnancies. Ninety-two cases of 214 suffered intrauterine demise (68 selective pregnancy terminations and 24 stillbirths) and 58 died in infancy. In 141 out of these cases the diagnosis could be included within lethal arthrogryposes, with a prevalence of 1 in 6,985 (1.43/10,000) births. Of these, 59 had spinal cord pathology at autopsy and thus were of neurogenic origin. Thirty-nine cases had lethal congenital contracture syndrome (LCCS) clinically characterized by total immobility of the fetus at all ultrasound examinations (12 weeks or later), multiple joint contractures in both upper and lower limbs, hydrops, and fetal death before the 32nd week of pregnancy. LCCS is noted as a unique Finnish disorder with a prevalence of 1 in 25,250 (0.40/10,000) births and is a major cause of lethal arthrogryposis in Finland.

Radioimmunoassay for yeast killer toxin from Saccharomyces cerevisiae

International Nuclear Information System (INIS)

Siddiqui, F.A.; Bussey, H.

1981-01-01

A radioimmunoassay was developed for the K1 killer toxin from strain T158C/S14a of Saccharomyces cerevisiae. Iodine 125-labelled toxin was made to a specific activity of 100 μCi/mg of protein. Antibody to purified toxin was prepared in rabbits using toxin cross-linked to itself. These antibodies, partially purified by 50 percent ammonium sulfate precipitation and Sepharose CL-6B column chromatography, produced one precipitation band with killer toxin and bound 125 I-labelled toxin in a radioimmunoassay. The antibody preparation also bound with the toxins from another K1 killer, A364A, and three chromosomal superkiller mutants derived from it. (auth)

The botulinum toxin as a therapeutic agent: molecular and pharmacological insights

Directory of Open Access Journals (Sweden)

Kukreja R

2015-12-01

Full Text Available Roshan Kukreja,1 Bal Ram Singh2 1Department of Chemistry and Biochemistry, University of Massachusetts, 2Botulinum Research Center, Institute of Advanced Sciences, Dartmouth, MA, USA Abstract: Botulinum neurotoxins (BoNTs, the most potent toxins known to mankind, are metalloproteases that act on nerve–muscle junctions to block exocytosis through a very specific and exclusive endopeptidase activity against soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE proteins of presynaptic vesicle fusion machinery. This very ability of the toxins to produce flaccid muscle paralysis through chemical denervation has been put to good use, and these potentially lethal toxins have been licensed to treat an ever expanding list of medical disorders and more popularly in the field of esthetic medicine. In most cases, therapeutic BoNT preparations are high-molecular-weight protein complexes consisting of BoNT, complexing proteins, and excipients. There is at least one isolated BoNT, which is free of complexing proteins in the market (Xeomin®. Each commercially available BoNT formulation is unique, differing mainly in molecular size and composition of complexing proteins, biological activity, and antigenicity. BoNT serotype A is marketed as Botox®, Dysport®, and Xeomin®, while BoNT type B is commercially available as Myobloc®. Nerve terminal intoxication by BoNTs is completely reversible, and the duration of therapeutic effects of BoNTs varies for different serotypes. Depending on the target tissue, BoNTs can block the cholinergic neuromuscular or cholinergic autonomic innervation of exocrine glands and smooth muscles. Therapeutic BoNTs exhibit a high safety and very limited adverse effects profile. Despite their established efficacy, the greatest concern with the use of therapeutic BoNTs is their propensity to elicit immunogenic reactions that might render the patient unresponsive to subsequent treatments, particularly in chronic

In vitro cell culture lethal dose submitted to gamma radiation

Energy Technology Data Exchange (ETDEWEB)

Moreno, Carolina S.; Rogero, Sizue O.; Rogero, Jose Roberto [Instituto de Pesquisas Energeticas e Nucleares (IPEN/CNEN-SP), Sao Paulo, SP (Brazil)], e-mail: carolina_sm@hotmail.com; Ikeda, Tamiko I.; Cruz, Aurea S. [Instituto Adolfo Lutz, Sao Paulo, SP (Brazil)

2009-07-01

The present study was designed to evaluate the in vitro effect of gamma radiation in cell culture of mouse connective tissue exposed to different doses of gamma radiation and under several conditions. The cell viability was analyzed by neutral red uptake methodology. This assay was developed for establish a methodology to be used in the future in the study of resveratrol radioprotection. Resveratrol (3,4',5- trihydroxystilbene), a phenolic phytoalexin that occurs naturally in some spermatophytes, such as grapevines, in response to injury as fungal infections and exposure to ultraviolet light. In the wines this compound is found at high levels and is considered one of the highest antioxidant constituents. The intense antioxidant potential of resveratrol provides many pharmacological activities including cardioprotection, chemoprevention and anti-tumor effects. Our results demonstrated that {sup 60}Co gamma radiation lethal dose (LD50) on NCTC clone 929 cells was about 340Gy. (author)

In vitro cell culture lethal dose submitted to gamma radiation

International Nuclear Information System (INIS)

Moreno, Carolina S.; Rogero, Sizue O.; Rogero, Jose Roberto; Ikeda, Tamiko I.; Cruz, Aurea S.

2009-01-01

The present study was designed to evaluate the in vitro effect of gamma radiation in cell culture of mouse connective tissue exposed to different doses of gamma radiation and under several conditions. The cell viability was analyzed by neutral red uptake methodology. This assay was developed for establish a methodology to be used in the future in the study of resveratrol radioprotection. Resveratrol (3,4',5- trihydroxystilbene), a phenolic phytoalexin that occurs naturally in some spermatophytes, such as grapevines, in response to injury as fungal infections and exposure to ultraviolet light. In the wines this compound is found at high levels and is considered one of the highest antioxidant constituents. The intense antioxidant potential of resveratrol provides many pharmacological activities including cardioprotection, chemoprevention and anti-tumor effects. Our results demonstrated that 60 Co gamma radiation lethal dose (LD50) on NCTC clone 929 cells was about 340Gy. (author)

Marine and freshwater toxins.

Science.gov (United States)

Hungerford, James M

2006-01-01

In a very busy and exciting year, 2005 included First Action approval of a much needed official method for paralytic shellfish toxins and multiple international toxin symposia highlighted by groundbreaking research. These are the first-year milestones and activities of the Marine and Freshwater Toxins Task Force and Analytical Community. Inaugurated in 2004 and described in detail in last year's General Referee Report (1) this international toxins group has grown to 150 members from many regions and countries. Perhaps most important they are now making important and global contributions to food safety and to providing alternatives to animal-based assays. Official Method 2005.06 was first approved in late 2004 by the Task Force and subsequently Official First Action in 2005 (2) by the Methods Committee on Natural Toxins and Food Allergens and the Official Methods Board. This nonproprietary method (3) is a precolumn oxidation, liquid chromatographic method that makes good use of fluorescence detection to provide high sensitivity detection of the saxitoxins. It has also proven to be rugged enough for regulatory use and the highest level of validation. As pointed out in the report of method principle investigator and Study Director James Lawrence, approval of 2005.06 now provides the first official alternative to the mouse bioassay after many decades of shellfish monitoring. This past year in April 2005 the group also held their first international conference, "Marine and Freshwater Toxins Analysis: Ist Joint Symposium and AOAC Task Force Meeting," in Baiona, Spain. The 4-day conference consisted of research and stakeholder presentations and symposium-integrated subgroup sessions on ciguatoxins, saxitoxin assays and liquid chromatography (LC) methods for saxitoxins and domoic acids, okadaiates and azaspiracids, and yessotoxins. Many of these subgroups were recently formed in 2005 and are working towards their goals of producing officially validated analytical methods

Identification of a human monoclonal antibody to replace equine diphtheria antitoxin for treatment of diphtheria intoxication.

Science.gov (United States)

Sevigny, Leila M; Booth, Brian J; Rowley, Kirk J; Leav, Brett A; Cheslock, Peter S; Garrity, Kerry A; Sloan, Susan E; Thomas, William; Babcock, Gregory J; Wang, Yang

2013-11-01

Diphtheria antitoxin (DAT) has been the cornerstone of the treatment of Corynebacterium diphtheriae infection for more than 100 years. Although the global incidence of diphtheria has declined steadily over the last quarter of the 20th century, the disease remains endemic in many parts of the world, and significant outbreaks still occur. DAT is an equine polyclonal antibody that is not commercially available in the United States and is in short supply globally. A safer, more readily available alternative to DAT would be desirable. In the current study, we obtained human monoclonal antibodies (hMAbs) directly from antibody-secreting cells in the circulation of immunized human volunteers. We isolated a panel of diverse hMAbs that recognized diphtheria toxoid, as well as a variety of recombinant protein fragments of diphtheria toxin. Forty-five unique hMAbs were tested for neutralization of diphtheria toxin in in vitro cytotoxicity assays with a 50% effective concentration of 0.65 ng/ml for the lead candidate hMAb, 315C4. In addition, 25 μg of 315C4 completely protected guinea pigs from intoxication in an in vivo lethality model, yielding an estimated relative potency of 64 IU/mg. In comparison, 1.6 IU of DAT was necessary for full protection from morbidity and mortality in this model. We further established that our lead candidate hMAb binds to the receptor-binding domain of diphtheria toxin and physically blocks the toxin from binding to the putative receptor, heparin-binding epidermal growth factor-like growth factor. The discovery of a specific and potent human neutralizing antibody against diphtheria toxin holds promise as a potential therapeutic.

Role of Botulinum Toxin in Depression.

Science.gov (United States)

Parsaik, Ajay K; Mascarenhas, Sonia S; Hashmi, Aqeel; Prokop, Larry J; John, Vineeth; Okusaga, Olaoluwa; Singh, Balwinder

2016-03-01

The goal of this review was to consolidate the evidence concerning the efficacy of botulinum toxin type A (onabotulinumtoxinA) in depression. We searched MEDLINE, EMBASE, Cochrane, and Scopus through May 5, 2014, for studies evaluating the efficacy of botulinum toxin A in depression. Only randomized controlled trials were included in the meta-analysis. A pooled mean difference in primary depression score, and pooled odds ratio for response and remission rate with 95% confidence interval (CI) were estimated using the random-effects model. Heterogeneity was assessed using Cochran Q test and χ statistic. Of the 639 articles that were initially retrieved, 5 studies enrolling 194 subjects (age 49±9.6 y) were included in the systematic review, and 3 randomized controlled trials enrolling 134 subjects were included in the meta-analysis. The meta-analysis showed a significant decrease in mean primary depression scores among patients who received botulinum toxin A compared with placebo (-9.80; 95% CI, -12.90 to -6.69) with modest heterogeneity between the studies (Cochran Q test, χ=70). Response and remission rates were 8.3 and 4.6 times higher, respectively, among patients receiving botulinum toxin A compared with placebo, with no heterogeneity between the studies. The 2 studies excluded from the meta-analysis also found a significant decrease in primary depression scores in patients after receiving botulinum toxin A. A few subjects had minor side effects, which were similar between the groups receiving botulinum toxin and those receiving placebo. This study suggests that botulinum toxin A can produce significant improvement in depressive symptoms and is a safe adjunctive treatment for patients receiving pharmacotherapy for depression. Future trials are needed to evaluate the antidepressant effect per se of botulinum toxin A and to further elucidate the underlying antidepressant mechanism of botulinum toxin A.

Botulinum toxin therapy for limb dystonias.

Science.gov (United States)

Yoshimura, D M; Aminoff, M J; Olney, R K

1992-03-01

We investigated the effectiveness of botulinum toxin in 17 patients with limb dystonias (10 with occupational cramps, three with idiopathic dystonia unrelated to activity, and two each with post-stroke and parkinsonian dystonia) in a placebo-controlled, blinded study. We identified affected muscles clinically and by recording the EMG from implanted wire electrodes at rest and during performance of tasks that precipitated abnormal postures. There were three injections given with graded doses of toxin (average doses, 5 to 10, 10 to 20, and 20 to 40 units per muscle) and one with placebo, in random order. Subjective improvement occurred after 53% of injections of botulinum toxin, and this was substantial in 24%. Only one patient (7%) improved after placebo injection. Subjective improvement occurred in 82% of patients with at least one dose of toxin, lasting for 1 to 4 months. Response rates were similar between clinical groups. Objective evaluation failed to demonstrate significant improvement following treatment with toxin compared with placebo. The major side effect was transient focal weakness after 53% of injections of toxin.

Fatal Clostridium botulinum toxicosis in eleven Holstein cattle fed round bale barley haylage.

Science.gov (United States)

Kelch, W J; Kerr, L A; Pringle, J K; Rohrbach, B W; Whitlock, R H

2000-09-01

Twenty-two lactating Holstein cattle in Tennessee had clinical signs of intoxication with preformed Clostridium botulinum toxin. These signs included weakness, paralysis of the tongue and chest muscles, abdominal breathing, and, in 11 of the 22 cows, death. Differential diagnoses included hypocalcemia, hypomagnesemia, carbohydrate overload, and several toxicoses including mycotoxin, lead, nitrate, organophosphate, atropine or atropine-like alkaloid, and botulism. A diagnosis of botulism by the ingestion of preformed C. botulinum type B toxin was made by eliminating these other diseases, by finding C. botulinum type B spores in 3 bales of round bale barley haylage fed to these cattle, and by isolating preformed type B toxin from 1 of the 3 bales. Confirmation of the toxin type was made by demonstrating mouse lethality by intraperitoneal injection of specimen extracts with neutralization by C. botulinum type B antitoxin. The haylage, harvested green and encased in black plastic bags to facilitate fermentation, was presumably contaminated by the botulinum toxin when fermentation failed to produce enough acid to lower the pH to 4.5, the pH below which C. botulinum growth is inhibited. Farmers and ranchers who use round hay balers to produce haylage should be alert to this potential problem.

Recent Insights into Clostridium perfringens Beta-Toxin

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Masahiro Nagahama

2015-02-01

Full Text Available Clostridium perfringens beta-toxin is a key mediator of necrotizing enterocolitis and enterotoxemia. It is a pore-forming toxin (PFT that exerts cytotoxic effect. Experimental investigation using piglet and rabbit intestinal loop models and a mouse infection model apparently showed that beta-toxin is the important pathogenic factor of the organisms. The toxin caused the swelling and disruption of HL-60 cells and formed a functional pore in the lipid raft microdomains of sensitive cells. These findings represent significant progress in the characterization of the toxin with knowledge on its biological features, mechanism of action and structure-function having been accumulated. Our aims here are to review the current progresses in our comprehension of the virulence of C. perfringens type C and the character, biological feature and structure-function of beta-toxin.

Neutralization of antibody-enhanced dengue infection by VIS513, a pan serotype reactive monoclonal antibody targeting domain III of the dengue E protein

Science.gov (United States)

Robinson, Luke N.; Ong, Li Ching; Rowley, Kirk J.; Winnett, Alexander; Tan, Hwee Cheng; Hobbie, Sven; Shriver, Zachary; Babcock, Gregory J.; Alonso, Sylvie; Ooi, Eng Eong

2018-01-01

Dengue virus (DENV) infection imposes enormous health and economic burden worldwide with no approved treatment. Several small molecules, including lovastatin, celgosivir, balapiravir and chloroquine have been tested for potential anti-dengue activity in clinical trials; none of these have demonstrated a protective effect. Recently, based on identification and characterization of cross-serotype neutralizing antibodies, there is increasing attention on the potential for dengue immunotherapy. Here, we tested the ability of VIS513, an engineered cross-neutralizing humanized antibody targeting the DENV E protein domain III, to overcome antibody-enhanced infection and high but brief viremia, which are commonly encountered in dengue patients, in various in vitro and in vivo models. We observed that VIS513 efficiently neutralizes DENV at clinically relevant viral loads or in the presence of enhancing levels of DENV immune sera. Single therapeutic administration of VIS513 in mouse models of primary infection or lethal secondary antibody-enhanced infection, reduces DENV titers and protects from lethal infection. Finally, VIS513 administration does not readily lead to resistance, either in cell culture systems or in animal models of dengue infection. The findings suggest that rapid viral reduction during acute DENV infection with a monoclonal antibody is feasible. PMID:29425203

Military Importance of Natural Toxins and Their Analogs

Directory of Open Access Journals (Sweden)

Vladimír Pitschmann

2016-04-01

Full Text Available Toxin weapon research, development, production and the ban on its uses is an integral part of international law, with particular attention paid to the protection against these weapons. In spite of this, hazards associated with toxins cannot be completely excluded. Some of these hazards are also pointed out in the present review. The article deals with the characteristics and properties of natural toxins and synthetic analogs potentially constituting the basis of toxin weapons. It briefly describes the history of military research and the use of toxins from distant history up to the present age. With respect to effective disarmament conventions, it mentions certain contemporary concepts of possible toxin applications for military purposes and the protection of public order (suppression of riots; it also briefly refers to the question of terrorism. In addition, it deals with certain traditional as well as modern technologies of the research, synthesis, and use of toxins, which can affect the continuing development of toxin weapons. These are, for example, cases of new toxins from natural sources, their chemical synthesis, production of synthetic analogs, the possibility of using methods of genetic engineering and modern biotechnologies or the possible applications of nanotechnology and certain pharmaceutical methods for the effective transfer of toxins into the organism. The authors evaluate the military importance of toxins based on their comparison with traditional chemical warfare agents. They appeal to the ethics of the scientific work as a principal condition for the prevention of toxin abuse in wars, military conflicts, as well as in non-military attacks.

Military Importance of Natural Toxins and Their Analogs.

Science.gov (United States)

Pitschmann, Vladimír; Hon, Zdeněk

2016-04-28

Toxin weapon research, development, production and the ban on its uses is an integral part of international law, with particular attention paid to the protection against these weapons. In spite of this, hazards associated with toxins cannot be completely excluded. Some of these hazards are also pointed out in the present review. The article deals with the characteristics and properties of natural toxins and synthetic analogs potentially constituting the basis of toxin weapons. It briefly describes the history of military research and the use of toxins from distant history up to the present age. With respect to effective disarmament conventions, it mentions certain contemporary concepts of possible toxin applications for military purposes and the protection of public order (suppression of riots); it also briefly refers to the question of terrorism. In addition, it deals with certain traditional as well as modern technologies of the research, synthesis, and use of toxins, which can affect the continuing development of toxin weapons. These are, for example, cases of new toxins from natural sources, their chemical synthesis, production of synthetic analogs, the possibility of using methods of genetic engineering and modern biotechnologies or the possible applications of nanotechnology and certain pharmaceutical methods for the effective transfer of toxins into the organism. The authors evaluate the military importance of toxins based on their comparison with traditional chemical warfare agents. They appeal to the ethics of the scientific work as a principal condition for the prevention of toxin abuse in wars, military conflicts, as well as in non-military attacks.

Chronic Exposure of Corals to Fine Sediments: Lethal and Sub-Lethal Impacts

Science.gov (United States)

Flores, Florita; Hoogenboom, Mia O.; Smith, Luke D.; Cooper, Timothy F.; Abrego, David; Negri, Andrew P.

2012-01-01

Understanding the sedimentation and turbidity thresholds for corals is critical in assessing the potential impacts of dredging projects in tropical marine systems. In this study, we exposed two species of coral sampled from offshore locations to six levels of total suspended solids (TSS) for 16 weeks in the laboratory, including a 4 week recovery period. Dose-response relationships were developed to quantify the lethal and sub-lethal thresholds of sedimentation and turbidity for the corals. The sediment treatments affected the horizontal foliaceous species (Montipora aequituberculata) more than the upright branching species (Acropora millepora). The lowest sediment treatments that caused full colony mortality were 30 mg l−1 TSS (25 mg cm−2 day−1) for M. aequituberculata and 100 mg l−1 TSS (83 mg cm−2 day−1) for A. millepora after 12 weeks. Coral mortality generally took longer than 4 weeks and was closely related to sediment accumulation on the surface of the corals. While measurements of damage to photosystem II in the symbionts and reductions in lipid content and growth indicated sub-lethal responses in surviving corals, the most reliable predictor of coral mortality in this experiment was long-term sediment accumulation on coral tissue. PMID:22662225

Chronic exposure of corals to fine sediments: lethal and sub-lethal impacts.

Directory of Open Access Journals (Sweden)

Florita Flores

Full Text Available Understanding the sedimentation and turbidity thresholds for corals is critical in assessing the potential impacts of dredging projects in tropical marine systems. In this study, we exposed two species of coral sampled from offshore locations to six levels of total suspended solids (TSS for 16 weeks in the laboratory, including a 4 week recovery period. Dose-response relationships were developed to quantify the lethal and sub-lethal thresholds of sedimentation and turbidity for the corals. The sediment treatments affected the horizontal foliaceous species (Montipora aequituberculata more than the upright branching species (Acropora millepora. The lowest sediment treatments that caused full colony mortality were 30 mg l(-1 TSS (25 mg cm(-2 day(-1 for M. aequituberculata and 100 mg l(-1 TSS (83 mg cm(-2 day(-1 for A. millepora after 12 weeks. Coral mortality generally took longer than 4 weeks and was closely related to sediment accumulation on the surface of the corals. While measurements of damage to photosystem II in the symbionts and reductions in lipid content and growth indicated sub-lethal responses in surviving corals, the most reliable predictor of coral mortality in this experiment was long-term sediment accumulation on coral tissue.

Toxins of filamentous fungi.

Science.gov (United States)

Bhatnagar, Deepak; Yu, Jiujiang; Ehrlich, Kenneth C

2002-01-01

Mycotoxins are low-molecular-weight secondary metabolites of fungi. The most significant mycotoxins are contaminants of agricultural commodities, foods and feeds. Fungi that produce these toxins do so both prior to harvest and during storage. Although contamination of commodities by toxigenic fungi occurs frequently in areas with a hot and humid climate (i.e. conditions favorable for fungal growth), they can also be found in temperate conditions. Production of mycotoxins is dependent upon the type of producing fungus and environmental conditions such as the substrate, water activity (moisture and relative humidity), duration of exposure to stress conditions and microbial, insect or other animal interactions. Although outbreaks of mycotoxicoses in humans have been documented, several of these have not been well characterized, neither has a direct correlation between the mycotoxin and resulting toxic effect been well established in vivo. Even though the specific modes of action of most of the toxins are not well established, acute and chronic effects in prokaryotic and eukaryotic systems, including humans have been reported. The toxicity of the mycotoxins varies considerably with the toxin, the animal species exposed to it, and the extent of exposure, age and nutritional status. Most of the toxic effects of mycotoxins are limited to specific organs, but several mycotoxins affect many organs. Induction of cancer by some mycotoxins is a major concern as a chronic effect of these toxins. It is nearly impossible to eliminate mycotoxins from the foods and feed in spite of the regulatory efforts at the national and international levels to remove the contaminated commodities. This is because mycotoxins are highly stable compounds, the producing fungi are ubiquitous, and food contamination can occur both before and after harvest. Nevertheless, good farm management practices and adequate storage facilities minimize the toxin contamination problems. Current research is

Effects of ionizing radiation on crotoxin (toxin of Crotalus durissus terrificus venom): molecular studies

International Nuclear Information System (INIS)

Souza Filho, J.N. de.

1988-01-01

It is know that the ionizing radiation is able to change significantly the biological and antigenic response of a toxin depending of the dose and irradiation's conditions, probable by structural alterations caused by radiation. In this work, the crotoxin, principal neurotoxin of the South American rattlesnake venom, was isolated using molecular exclusion chromatography with Sephadex G-75 and follwed by precipitation on the isoelectric point. Fractions in the concentration of 2 mg of protein/m1 0.85% NaCl were irradiated in a source of sup(60)Co GAMMACELL with dose rate of 1100 Gy/h using doses of 250, 500, 1000, 1500 and 2000 Gy. It was determinated for these samples, the proteic concentration (Lowry's method), the content sulphydryl (Ellman's method), the profile electrophoretic (SDS-PAGE), the toxicity by lethal dose 50% in mice and the antigenic response using crotalic antiserum by the diffusion imunoassay (Ouchterlony's method). The results showed the formation of aggregates and loss of protein in solution by precipitation. In the dose of 1000 Gy and higher it was possible to observe the presence of sulphydryl groups indicating the breakage of S-S bridges. The lethal dose 50% increased 2 times for the dose of 1000 Gy and 3.5 times for 1500 Gy shoding a detoxication. By the other hand, the antigenic response seems to be still intact at doses up to 1000 Gy. (author)

Toxin-Antitoxin Battle in Bacteria

DEFF Research Database (Denmark)

Cataudella, Ilaria

This PhD thesis consists of three research projects revolving around the common thread of investigation of the properties and biological functions of Toxin-Antitoxin loci. Toxin-Antitoxin (TA) loci are transcriptionally regulated via an auto-inhibition mechanism called conditional cooperativity, ...

Bio Warfare and Terrorism: Toxins and Other Mid-Spectrum Agents

National Research Council Canada - National Science Library

Madsen, James M

2005-01-01

... counterparts are still by definition toxins. Related terms include phycotoxins (toxins from algae), mycotoxins (fungal toxins), phytotoxins (plant toxins), and venoms (toxins from animals, especially vertebrates...

Engineering toxins for 21st century therapies.

Science.gov (United States)

Chaddock, John A; Acharya, K Ravi

2011-04-01

'Engineering Toxins for 21st Century Therapies' (9-10 September 2010) was part of the Royal Society International Seminar series held at the Kavli International Centre, UK. Participants were assembled from a range of disciplines (academic, industry, regulatory, public health) to discuss the future potential of toxin-based therapies. The meeting explored how the current structural and mechanistic knowledge of toxins could be used to engineer future toxin-based therapies. To date, significant progress has been made in the design of novel recombinant biologics based on domains of natural toxins, engineered to exhibit advantageous properties. The meeting concluded, firstly that future product development vitally required the appropriate combination of creativity and innovation that can come from the academic, biotechnology and pharma sectors. Second, that continued investigation into understanding the basic science of the toxins and their targets was essential in order to develop new opportunities for the existing products and to create new products with enhanced properties. Finally, it was concluded that the clinical potential for development of novel biologics based on toxin domains was evident. © 2011 The Authors Journal compilation © 2011 FEBS.

Crystallization of isoelectrically homogeneous cholera toxin

International Nuclear Information System (INIS)

Spangler, B.D.; Westbrook, E.M.

1989-01-01

Past difficulty in growing good crystals of cholera toxin has prevented the study of the crystal structure of this important protein. The authors have determined that failure of cholera toxin to crystallize well has been due to its heterogeneity. They have now succeeded in overcoming the problem by isolating a single isoelectric variant of this oligomeric protein (one A subunit and five B subunits). Cholera toxin purified by their procedure readily forms large single crystals. The crystal form has been described previously. They have recorded data from native crystals of cholera toxin to 3.0-angstrom resolution with our electronic area detectors. With these data, they have found the orientation of a 5-fold symmetry axis within these crystals, perpendicular to the screw dyad of the crystal. They are now determining the crystal structure of cholera toxin by a combination of multiple heavy-atom isomorphous replacement and density modification techniques, making use of rotational 5-fold averaging of the B subunits

Botulinum toxin in trigeminal neuralgia.

Science.gov (United States)

Castillo-Álvarez, Federico; Hernando de la Bárcena, Ignacio; Marzo-Sola, María Eugenia

2017-01-06

Trigeminal neuralgia is one of the most disabling facial pain syndromes, with a significant impact on patients' quality of life. Pharmacotherapy is the first choice for treatment but cases of drug resistance often require new strategies, among which various interventional treatments have been used. In recent years a new therapeutic strategy consisting of botulinum toxin has emerged, with promising results. We reviewed clinical cases and case series, open-label studies and randomized clinical trials examining the use of botulinum toxin for drug-refractory trigeminal neuralgia published in the literature. The administration of botulinum toxin has proven to be a safe and effective therapeutic strategy in patients with drug-refractory idiopathic trigeminal neuralgia, but many questions remain unanswered as to the precise role of botulinum toxin in the treatment of this disease. Copyright © 2016 Elsevier España, S.L.U. All rights reserved.

Collaborative Research Program on Seafood Toxins

Science.gov (United States)

1988-08-14

Crystallographic Structures of Saxitoxins Cl and C2 Appendix C: Collaborative Research Program an Seafcod Toxins Progress Report on Ciguatera and Related...radioimmunoassay for PSP were also evalumted. The Hokama stick test for ciguatera toxin was also evaluated. 4. initiate Studies on the Accumulation...tco•d which caie a form of b-mnn poisoning referred to as ciguatera . The respcnsible toxins originate from ll1ular rine algae of the division

Comparison between IgG and F(ab′)2 polyvalent antivenoms: neutralization of systemic effects induced by Bothrops asper venom in mice, extravasation to muscle tissue, and potential for induction of adverse reactions

OpenAIRE

León Montero, Guillermo; Monge Monge, María; Rojas Umaña, Ermila; Lomonte, Bruno; Gutiérrez, José María

2001-01-01

Whole IgG and F(ab′)2 equine-derived polyvalent (Crotalinae) antivenoms, prepared from the same batch of hyperimmune plasma, were compared in terms of neutralization of the lethal and defibrinating activities induced by Bothrops asper venom, their ability to reach the muscle tissue compartment in envenomated mice, and their potential for the induction of adverse reactions. Both preparations were adjusted to the same potency against the lethal effect of B. asper venom in experiments involving ...

Entry of Shiga toxin into cells

DEFF Research Database (Denmark)

Sandvig, Kirsten; van Deurs, Bo

1994-01-01

Cellebiologi, Shiga toxin, receptors, glycolipids, endocytosis, trans-Golgi network, endoplasmic reticulum, retrograde transport......Cellebiologi, Shiga toxin, receptors, glycolipids, endocytosis, trans-Golgi network, endoplasmic reticulum, retrograde transport...

Botulinum Toxin: Pharmacology and Therapeutic Roles in Pain States.

Science.gov (United States)

Patil, Shilpadevi; Willett, Olga; Thompkins, Terin; Hermann, Robert; Ramanathan, Sathish; Cornett, Elyse M; Fox, Charles J; Kaye, Alan David

2016-03-01

Botulinum toxin, also known as Botox, is produced by Clostridium botulinum, a gram-positive anaerobic bacterium, and botulinum toxin injections are among the most commonly practiced cosmetic procedures in the USA. Although botulinum toxin is typically associated with cosmetic procedures, it can be used to treat a variety of other conditions, including pain. Botulinum toxin blocks the release of acetylcholine from nerve endings to paralyze muscles and to decrease the pain response. Botulinum toxin has a long duration of action, lasting up to 5 months after initial treatment which makes it an excellent treatment for chronic pain patients. This manuscript will outline in detail why botulinum toxin is used as a successful treatment for pain in multiple conditions as well as outline the risks associated with using botulinum toxin in certain individuals. As of today, the only FDA-approved chronic condition that botulinum toxin can be used to treat is migraines and this is related to its ability to decrease muscle tension and increase muscle relaxation. Contraindications to botulinum toxin treatments are limited to a hypersensitivity to the toxin or an infection at the site of injection, and there are no known drug interactions with botulinum toxin. Botulinum toxin is an advantageous and effective alternative pain treatment and a therapy to consider for those that do not respond to opioid treatment. In summary, botulinum toxin is a relatively safe and effective treatment for individuals with certain pain conditions, including migraines. More research is warranted to elucidate chronic and long-term implications of botulinum toxin treatment as well as effects in pregnant, elderly, and adolescent patients.

Loading and Light Degradation Characteristics of B t Toxin on Nano goethite: A Potential Material for Controlling the Environmental Risk of B t Toxin

International Nuclear Information System (INIS)

Zhou, X.; She, Ch.; She, Ch.; Liu, H.

2015-01-01

Transgenic B t-modified crops release toxins into soil through root exudate s and upon decomposition of residues. The fate of these toxins in soil has not been yet clearly elucidated. Nano goethite was found to have a different influence on the lifetime and identicalness activity of B t toxin. The aim of this study was to elucidate the adsorption characteristics of B t toxin on nano goethite and its activity changes before and after adsorption. The adsorption of toxin on nano goethite reached equilibrium within 5 h, and the adsorption isotherm of B t toxin on nano goethite conformed to the Langmuir equation (). In the range of ph from 6.0 to 8.0, larger adsorption occurred at lower ph value. The toxin adsorption decreased with the temperature between 10 and 50 degree. The results of Ftir, XRD, and SEM indicated that toxin did not influence the structure of nano goethite and the adsorption of toxin only on the surface of nano goethite. The LC_5_0 value for bound toxin was higher than that of free toxin, and the nano goethite greatly accelerated the degradation of toxin by ultraviolet irradiation. The above results suggested that nano goethite is a potential material for controlling the environmental risk of toxin released by Bt transgenic plants

Monoclonal antibodies and toxins--a perspective on function and isotype.

Science.gov (United States)

Chow, Siu-Kei; Casadevall, Arturo

2012-06-01

Antibody therapy remains the only effective treatment for toxin-mediated diseases. The development of hybridoma technology has allowed the isolation of monoclonal antibodies (mAbs) with high specificity and defined properties, and numerous mAbs have been purified and characterized for their protective efficacy against different toxins. This review summarizes the mAb studies for 6 toxins--Shiga toxin, pertussis toxin, anthrax toxin, ricin toxin, botulinum toxin, and Staphylococcal enterotoxin B (SEB)--and analyzes the prevalence of mAb functions and their isotypes. Here we show that most toxin-binding mAbs resulted from immunization are non-protective and that mAbs with potential therapeutic use are preferably characterized. Various common practices and caveats of protection studies are discussed, with the goal of providing insights for the design of future research on antibody-toxin interactions.

Computational Studies of Snake Venom Toxins

OpenAIRE

Paola G. Ojeda; David Ramírez; Jans Alzate-Morales; Julio Caballero; Quentin Kaas; Wendy González

2017-01-01

Most snake venom toxins are proteins, and participate to envenomation through a diverse array of bioactivities, such as bleeding, inflammation, and pain, cytotoxic, cardiotoxic or neurotoxic effects. The venom of a single snake species contains hundreds of toxins, and the venoms of the 725 species of venomous snakes represent a large pool of potentially bioactive proteins. Despite considerable discovery efforts, most of the snake venom toxins are still uncharacterized. Modern bioinformatics t...

Clostridium botulinum C2 toxin--new insights into the cellular up-take of the actin-ADP-ribosylating toxin.

Science.gov (United States)

Aktories, Klaus; Barth, Holger

2004-04-01

Clostridium botulinum C2 toxin is a member of the family of binary actin-ADP-ribosylating toxins. It consists of the enzyme component C2I, and the separated binding/translocation component C2II. Proteolytically activated C2II forms heptamers and binds to a carbohydrate cell surface receptor. After attachment of C2I, the toxin complex is endocytosed to reach early endosomes. At low pH of endosomes, C2II-heptamers insert into the membrane, form pores and deliver C2I into the cytosol. Here, C2I ADP-ribosylates actin at Arg177 to block actin polymerization and to induce depolymerization of actin filaments. The mini-review describes main properties of C2 toxin and discusses new findings on the involvement of chaperones in the up-take process of the toxin.

Toxins That Affect Voltage-Gated Sodium Channels.

Science.gov (United States)

Ji, Yonghua

2017-10-26

Voltage-gated sodium channels (VGSCs) are critical in generation and conduction of electrical signals in multiple excitable tissues. Natural toxins, produced by animal, plant, and microorganisms, target VGSCs through diverse strategies developed over millions of years of evolutions. Studying of the diverse interaction between VGSC and VGSC-targeting toxins has been contributing to the increasing understanding of molecular structure and function, pharmacology, and drug development potential of VGSCs. This chapter aims to summarize some of the current views on the VGSC-toxin interaction based on the established receptor sites of VGSC for natural toxins.

Cyanobacterial toxins: risk management for health protection

International Nuclear Information System (INIS)

Codd, Geoffrey A.; Morrison, Louise F.; Metcalf, James S.

2005-01-01

This paper reviews the occurrence and properties of cyanobacterial toxins, with reference to the recognition and management of the human health risks which they may present. Mass populations of toxin-producing cyanobacteria in natural and controlled waterbodies include blooms and scums of planktonic species, and mats and biofilms of benthic species. Toxic cyanobacterial populations have been reported in freshwaters in over 45 countries, and in numerous brackish, coastal, and marine environments. The principal toxigenic genera are listed. Known sources of the families of cyanobacterial toxins (hepato-, neuro-, and cytotoxins, irritants, and gastrointestinal toxins) are briefly discussed. Key procedures in the risk management of cyanobacterial toxins and cells are reviewed, including derivations (where sufficient data are available) of tolerable daily intakes (TDIs) and guideline values (GVs) with reference to the toxins in drinking water, and guideline levels for toxigenic cyanobacteria in bathing waters. Uncertainties and some gaps in knowledge are also discussed, including the importance of exposure media (animal and plant foods), in addition to potable and recreational waters. Finally, we present an outline of steps to develop and implement risk management strategies for cyanobacterial cells and toxins in waterbodies, with recent applications and the integration of Hazard Assessment Critical Control Point (HACCP) principles

Botulinum toxin for the treatment of bruxism.

Science.gov (United States)

Tinastepe, Neslihan; Küçük, Burcu Bal; Oral, Koray

2015-10-01

Botulinum toxin, the most potent biological toxin, has been shown to be effective for a variety of disorders in several medical conditions, when used both therapeutically and cosmetically. In recent years, there has been a rising trend in the use of this pharmacological agent to control bruxing activity, despite its reported adverse effects. The aim of this review was to provide a brief overview to clarify the underlying essential ideas for the use of botulinum toxin in bruxism based on available scientific papers. An electronic literature search was performed to identify publications related to botulinum toxin and its use for bruxism in PubMed. Hand searching of relevant articles was also made to identify additional studies. Of the eleven identified studies, only two were randomized controlled trials, compared with the effectiveness of botulinum toxins on the reduction in the frequency of bruxism events and myofascial pain after injection. The authors of these studies concluded that botulinum toxin could be used as an effective treatment for reducing nocturnal bruxism and myofascial pain in patients with bruxism. Evidence-based research was limited on this topic. More randomized controlled studies are needed to confirm that botulinum toxin is safe and reliable for routine clinical use in bruxism.

Mining of lethal recessive genetic variation in Danish cattle

DEFF Research Database (Denmark)

Das, Ashutosh

2015-01-01

in fertility. The primary objective of this PhD projekt was to identify recessive lethal gentic variants in the main Danish dairy cattle breed. Holstein-Friesian utilzing next generation sequencing (NGS) data. This study shows a potential for the use of the NGS-based reverse genetic approach in identifying...... lethal or semi-lethal recessive gentic variation...

Binding of Diphtheria Toxin to Phospholipids in Liposomes

Science.gov (United States)

Alving, Carl R.; Iglewski, Barbara H.; Urban, Katharine A.; Moss, Joel; Richards, Roberta L.; Sadoff, Jerald C.

1980-04-01

Diphtheria toxin bound to the phosphate portion of some, but not all, phospholipids in liposomes. Liposomes consisting of dimyristoyl phosphatidylcholine and cholesterol did not bind toxin. Addition of 20 mol% (compared to dimyristoyl phosphatidylcholine) of dipalmitoyl phosphatidic acid, dicetyl phosphate, phosphatidylinositol phosphate, cardiolipin, or phosphatidylserine in the liposomes resulted in substantial binding of toxin. Inclusion of phosphatidylinositol in dimyristol phosphatidylcholine / cholesterol liposomes did not result in toxin binding. The calcium salt of dipalmitoyl phosphatidic acid was more effective than the sodium salt, and the highest level of binding occurred with liposomes consisting only of dipalmitoyl phosphatidic acid (calcium salt) and cholesterol. Binding of toxin to liposomes was dependent on pH, and the pattern of pH dependence varied with liposomes having different compositions. Incubation of diphtheria toxin with liposomes containing dicetyl phosphate resulted in maximal binding at pH 3.6, whereas binding to liposomes containing phosphatidylinositol phosphate was maximal above pH 7. Toxin did not bind to liposomes containing 20 mol% of a free fatty acid (palmitic acid) or a sulfated lipid (3-sulfogalactosylceramide). Toxin binding to dicetyl phosphate or phosphatidylinositol phosphate was inhibited by UTP, ATP, phosphocholine, or p-nitrophenyl phosphate, but not by uracil. We conclude that (a) diphtheria toxin binds specifically to the phosphate portion of certain phospholipids, (b) binding to phospholipids in liposomes is dependent on pH, but is not due only to electrostatic interaction, and (c) binding may be strongly influenced by the composition of adjacent phospholipids that do not bind toxin. We propose that a minor membrane phospholipid (such as phosphatidylinositol phosphate or phosphatidic acid), or that some other phosphorylated membrane molecule (such as a phosphoprotein) may be important in the initial binding of

Can a toxin gene NAAT be used to predict toxin EIA and the severity of Clostridium difficile infection?

Directory of Open Access Journals (Sweden)

Mark I. Garvey

2017-12-01

Full Text Available Abstract Background Diagnosis of C. difficile infection (CDI is controversial because of the many laboratory methods available and their lack of ability to distinguish between carriage, mild or severe disease. Here we describe whether a low C. difficile toxin B nucleic acid amplification test (NAAT cycle threshold (CT can predict toxin EIA, CDI severity and mortality. Methods A three-stage algorithm was employed for CDI testing, comprising a screening test for glutamate dehydrogenase (GDH, followed by a NAAT, then a toxin enzyme immunoassay (EIA. All diarrhoeal samples positive for GDH and NAAT between 2012 and 2016 were analysed. The performance of the NAAT CT value as a classifier of toxin EIA outcome was analysed using a ROC curve; patient mortality was compared to CTs and toxin EIA via linear regression models. Results A CT value ≤26 was associated with ≥72% toxin EIA positivity; applying a logistic regression model we demonstrated an association between low CT values and toxin EIA positivity. A CT value of ≤26 was significantly associated (p = 0.0262 with increased one month mortality, severe cases of CDI or failure of first line treatment. The ROC curve probabilities demonstrated a CT cut off value of 26.6. Discussions Here we demonstrate that a CT ≤26 indicates more severe CDI and is associated with higher mortality. Samples with a low CT value are often toxin EIA positive, questioning the need for this additional EIA test. Conclusions A CT ≤26 could be used to assess the potential for severity of CDI and guide patient treatment.

Drooling in Parkinson's disease: A randomized controlled trial of incobotulinum toxin A and meta-analysis of Botulinum toxins.

Science.gov (United States)

Narayanaswami, Pushpa; Geisbush, Thomas; Tarulli, Andrew; Raynor, Elizabeth; Gautam, Shiva; Tarsy, Daniel; Gronseth, Gary

2016-09-01

Botulinum toxins are a therapeutic option for drooling in Parkinson's Disease (PD). The aims of this study were to: 1. evaluate the efficacy of incobotulinum toxin A for drooling in PD. 2. Perform a meta-analysis of studies of Botulinum toxins for drooling in PD. 1. Primary study: Randomized, double blind, placebo controlled, cross over trial. Incobotulinum toxin (100 units) or saline was injected into the parotid (20 units) and submandibular (30 units) glands. Subjects returned monthly for three evaluations after each injection. Outcome measures were saliva weight and Drooling Frequency and Severity Scale. 2. Systematic review of literature, followed by inverse variance meta-analyses using random effects models. 1. Primary Study: Nine of 10 subjects completed both arms. There was no significant change in the primary outcome of saliva weight one month after injection in the treatment period compared to placebo period (mean difference, gm ± SD: -0.194 ± 0.61, range: -1.28 to 0.97, 95% CI -0.71 to 0.32). Secondary outcomes also did not change. 2. Meta-analysis of six studies demonstrated significant benefit of Botulinum toxin on functional outcomes (effect size, Cohen's d: -1.32, CI -1.86 to -0.78). The other studies used a higher dose of Botulinum toxin A into the parotid glands. This study did not demonstrate efficacy of incobotulinum toxin A for drooling in PD, but lacked precision to exclude moderate benefit. The parotid/submandibular dose-ratio may have influenced results. Studies evaluating higher doses of incobotulinum toxin A into the parotid glands may be useful. Copyright © 2016 Elsevier Ltd. All rights reserved.

Array biosensor for detection of toxins

Science.gov (United States)

Ligler, Frances S.; Taitt, Chris Rowe; Shriver-Lake, Lisa C.; Sapsford, Kim E.; Shubin, Yura; Golden, Joel P.

2003-01-01

The array biosensor is capable of detecting multiple targets rapidly and simultaneously on the surface of a single waveguide. Sandwich and competitive fluoroimmunoassays have been developed to detect high and low molecular weight toxins, respectively, in complex samples. Recognition molecules (usually antibodies) were first immobilized in specific locations on the waveguide and the resultant patterned array was used to interrogate up to 12 different samples for the presence of multiple different analytes. Upon binding of a fluorescent analyte or fluorescent immunocomplex, the pattern of fluorescent spots was detected using a CCD camera. Automated image analysis was used to determine a mean fluorescence value for each assay spot and to subtract the local background signal. The location of the spot and its mean fluorescence value were used to determine the toxin identity and concentration. Toxins were measured in clinical fluids, environmental samples and foods, with minimal sample preparation. Results are shown for rapid analyses of staphylococcal enterotoxin B, ricin, cholera toxin, botulinum toxoids, trinitrotoluene, and the mycotoxin fumonisin. Toxins were detected at levels as low as 0.5 ng mL(-1).

Botulinum toxin for the treatment of strabismus.

Science.gov (United States)

Rowe, Fiona J; Noonan, Carmel P

2017-03-02

The use of botulinum toxin as an investigative and treatment modality for strabismus is well reported in the medical literature. However, it is unclear how effective it is in comparison to other treatment options for strabismus. The primary objective was to examine the efficacy of botulinum toxin therapy in the treatment of strabismus compared with alternative conservative or surgical treatment options. This review sought to ascertain those types of strabismus that particularly benefit from the use of botulinum toxin as a treatment option (such as small angle strabismus or strabismus with binocular potential, i.e. the potential to use both eyes together as a pair). The secondary objectives were to investigate the dose effect and complication rates associated with botulinum toxin. We searched CENTRAL (which contains the Cochrane Eyes and Vision Trials Register) (2016, Issue 6), Ovid MEDLINE, Ovid MEDLINE In-Process and Other Non-Indexed Citations, Ovid MEDLINE Daily, Ovid OLDMEDLINE (January 1946 to July 2016), Embase (January 1980 to July 2016), Latin American and Caribbean Literature on Health Sciences (LILACS) (January 1982 to July 2016), the ISRCTN registry (www.isrctn.com/editAdvancedSearch), ClinicalTrials.gov (www.clinicaltrials.gov), and the World Health Organization (WHO) International Clinical Trials Registry Platform (ICTRP) (www.who.int/ictrp/search/en). We did not use any date or language restrictions in the electronic searches for trials. We last searched the electronic databases on 11 July 2016. We handsearched the British and Irish Orthoptic Journal, Australian Orthoptic Journal, proceedings of the European Strabismological Association (ESA), International Strabismological Association (ISA) and International Orthoptic Association (IOA) (www.liv.ac.uk/orthoptics/research/search.htm) and American Academy of Paediatric Ophthalmology and Strabismus meetings (AAPOS). We contacted researchers who are active in this field for information about further

Human Neutralizing Monoclonal Antibody Inhibition of Middle East Respiratory Syndrome Coronavirus Replication in the Common Marmoset.

Science.gov (United States)

Chen, Zhe; Bao, Linlin; Chen, Cong; Zou, Tingting; Xue, Ying; Li, Fengdi; Lv, Qi; Gu, Songzhi; Gao, Xiaopan; Cui, Sheng; Wang, Jianmin; Qin, Chuan; Jin, Qi

2017-06-15

Middle East respiratory syndrome coronavirus (MERS-CoV) infection in humans is highly lethal, with a fatality rate of 35%. New prophylactic and therapeutic strategies to combat human infections are urgently needed. We isolated a fully human neutralizing antibody, MCA1, from a human survivor. The antibody recognizes the receptor-binding domain of MERS-CoV S glycoprotein and interferes with the interaction between viral S and the human cellular receptor human dipeptidyl peptidase 4 (DPP4). To our knowledge, this study is the first to report a human neutralizing monoclonal antibody that completely inhibits MERS-CoV replication in common marmosets. Monotherapy with MCA1 represents a potential alternative treatment for human infections with MERS-CoV worthy of evaluation in clinical settings. © Crown copyright 2017.

Single toxin dose-response models revisited

Energy Technology Data Exchange (ETDEWEB)

Demidenko, Eugene, E-mail: eugened@dartmouth.edu [Department of Biomedical Data Science, Geisel School of Medicine at Dartmouth, Hanover, NH03756 (United States); Glaholt, SP, E-mail: sglaholt@indiana.edu [Indiana University, School of Public & Environmental Affairs, Bloomington, IN47405 (United States); Department of Biological Sciences, Dartmouth College, Hanover, NH03755 (United States); Kyker-Snowman, E, E-mail: ek2002@wildcats.unh.edu [Department of Natural Resources and the Environment, University of New Hampshire, Durham, NH03824 (United States); Shaw, JR, E-mail: joeshaw@indiana.edu [Indiana University, School of Public & Environmental Affairs, Bloomington, IN47405 (United States); Chen, CY, E-mail: Celia.Y.Chen@dartmouth.edu [Department of Biological Sciences, Dartmouth College, Hanover, NH03755 (United States)

2017-01-01

The goal of this paper is to offer a rigorous analysis of the sigmoid shape single toxin dose-response relationship. The toxin efficacy function is introduced and four special points, including maximum toxin efficacy and inflection points, on the dose-response curve are defined. The special points define three phases of the toxin effect on mortality: (1) toxin concentrations smaller than the first inflection point or (2) larger then the second inflection point imply low mortality rate, and (3) concentrations between the first and the second inflection points imply high mortality rate. Probabilistic interpretation and mathematical analysis for each of the four models, Hill, logit, probit, and Weibull is provided. Two general model extensions are introduced: (1) the multi-target hit model that accounts for the existence of several vital receptors affected by the toxin, and (2) model with a nonzero mortality at zero concentration to account for natural mortality. Special attention is given to statistical estimation in the framework of the generalized linear model with the binomial dependent variable as the mortality count in each experiment, contrary to the widespread nonlinear regression treating the mortality rate as continuous variable. The models are illustrated using standard EPA Daphnia acute (48 h) toxicity tests with mortality as a function of NiCl or CuSO{sub 4} toxin. - Highlights: • The paper offers a rigorous study of a sigmoid dose-response relationship. • The concentration with highest mortality rate is rigorously defined. • A table with four special points for five morality curves is presented. • Two new sigmoid dose-response models have been introduced. • The generalized linear model is advocated for estimation of sigmoid dose-response relationship.

Interplay between toxin transport and flotillin localization

DEFF Research Database (Denmark)

Pust, Sascha; Dyve, Anne Berit; Torgersen, Maria L

2010-01-01

The flotillin proteins are localized in lipid domains at the plasma membrane as well as in intracellular compartments. In the present study, we examined the importance of flotillin-1 and flotillin-2 for the uptake and transport of the bacterial Shiga toxin (Stx) and the plant toxin ricin and we...... for flotillin-1 or -2. However, the Golgi-dependent sulfation of both toxins was significantly reduced in flotillin knockdown cells. Interestingly, when the transport of ricin to the ER was investigated, we obtained an increased mannosylation of ricin in flotillin-1 and flotillin-2 knockdown cells. The toxicity...... of both toxins was twofold increased in flotillin-depleted cells. Since BFA (Brefeldin A) inhibits the toxicity even in flotillin knockdown cells, the retrograde toxin transport is apparently still Golgi-dependent. Thus, flotillin proteins regulate and facilitate the retrograde transport of Stx and ricin....

Cholera Toxin B: One Subunit with Many Pharmaceutical Applications

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Keegan J. Baldauf

2015-03-01

Full Text Available Cholera, a waterborne acute diarrheal disease caused by Vibrio cholerae, remains prevalent in underdeveloped countries and is a serious health threat to those living in unsanitary conditions. The major virulence factor is cholera toxin (CT, which consists of two subunits: the A subunit (CTA and the B subunit (CTB. CTB is a 55 kD homopentameric, non-toxic protein binding to the GM1 ganglioside on mammalian cells with high affinity. Currently, recombinantly produced CTB is used as a component of an internationally licensed oral cholera vaccine, as the protein induces potent humoral immunity that can neutralize CT in the gut. Additionally, recent studies have revealed that CTB administration leads to the induction of anti-inflammatory mechanisms in vivo. This review will cover the potential of CTB as an immunomodulatory and anti-inflammatory agent. We will also summarize various recombinant expression systems available for recombinant CTB bioproduction.

The Influence of the Toxin/Antitoxin mazEF on Growth and Survival of Listeria monocytogenes under Stress

DEFF Research Database (Denmark)

Curtis, Thomas; Takeuchi, Ippei; Gram, Lone

2017-01-01

A major factor in the resilience of Listeria monocytogenes is the alternative sigma factor B (σB). Type II Toxin/Antitoxin (TA) systems are also known to have a role in the bacterial stress response upon activation via the ClpP or Lon proteases. Directly upstream of the σB operon in L....... monocytogenes is the TA system mazEF, which can cleave mRNA at UACMU sites. In this study, we showed that the mazEF TA locus does not affect the level of persister formation during treatment with antibiotics in lethal doses, but exerts different effects according to the sub-inhibitory stress added. Growth...... it is not analogous to the system of S. aureus, suggesting a novel mode of action for MazEF in L. monocytogenes....

Botulinum toxin for vaginismus treatment.

Science.gov (United States)

Ferreira, Juliana Rocha; Souza, Renan Pedra

2012-01-01

Vaginismus is characterized by recurrent or persistent involuntary contraction of the perineal muscles surrounding the outer third of the vagina when penile, finger, tampon, or speculum penetration is attempted. Recent results have suggested the use of botulinum toxin for the treatment of vaginismus. Here, we assessed previously published data to evaluate the therapeutic effectiveness of botulinum toxin for vaginismus. We have carried out a systematic review followed by a meta-analysis. Our results indicate that botulinum toxin is an effective therapeutic option for patients with vaginismus (pooled odds ratio of 8.723 with 95% confidence interval limits of 1.942 and 39.162, p = 0.005). This may hold particularly true in treatment-refractory patients because most of the studies included in this meta-analysis have enrolled these subjects in their primary analysis. Botulinum toxin appears to bea reasonable intervention for vaginismus. However, this conclusion should be read carefully because of the deficiency of placebo-controlled randomized clinical trials and the quality issues presented in the existing ones.

Use of the mice passive protection test to evaluate the humoral response in goats vaccinated with Sterne 34F2 live spore vaccine.

Science.gov (United States)

Phaswana, P H; Ndumnego, O C; Koehler, S M; Beyer, W; Crafford, J E; van Heerden, H

2017-09-07

The Sterne live spore vaccine (34F2) is the most widely used veterinary vaccine against anthrax in animals. Antibody responses to several antigens of Bacillus anthracis have been described with a large focus on those against protective antigen (PA). The focus of this study was to evaluate the protective humoral immune response induced by the live spore anthrax vaccine in goats. Boer goats vaccinated twice (week 0 and week 12) with the Sterne live spore vaccine and naive goats were used to monitor the anti-PA and toxin neutralizing antibodies at week 4 and week 17 (after the second vaccine dose) post vaccination. A/J mice were passively immunized with different dilutions of sera from immune and naive goats and then challenged with spores of B. anthracis strain 34F2 to determine the protective capacity of the goat sera. The goat anti-PA ELISA titres indicated significant sero-conversion at week 17 after the second doses of vaccine (p = 0.009). Mice receiving undiluted sera from goats given two doses of vaccine (twice immunized) showed the highest protection (86%) with only 20% of mice receiving 1:1000 diluted sera surviving lethal challenge. The in vitro toxin neutralization assay (TNA) titres correlated to protection of passively immunized A/J mice against lethal infection with the vaccine strain Sterne 34F2 spores using immune goat sera up to a 1:10 dilution (r s  ≥ 0.522, p = 0.046). This study suggests that the passive mouse protection model could be potentially used to evaluate the protective immune response in livestock animals vaccinated with the current live vaccine and new vaccines.

The involvement of IL-17A in the murine response to sub-lethal inhalational infection with Francisella tularensis.

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Gal Markel

2010-06-01

Full Text Available Francisella tularensis is an intercellular bacterium often causing fatal disease when inhaled. Previous reports have underlined the role of cell-mediated immunity and IFNgamma in the host response to Francisella tularensis infection.Here we provide evidence for the involvement of IL-17A in host defense to inhalational tularemia, using a mouse model of intranasal infection with the Live Vaccine Strain (LVS. We demonstrate the kinetics of IL-17A production in lavage fluids of infected lungs and identify the IL-17A-producing lymphocytes as pulmonary gammadelta and Th17 cells. The peak of IL-17A production appears early during sub-lethal infection, it precedes the peak of immune activation and the nadir of the disease, and then subsides subsequently. Exogenous airway administration of IL-17A or of IL-23 had a limited yet consistent effect of delaying the onset of death from a lethal dose of LVS, implying that IL-17A may be involved in restraining the infection. The protective role for IL-17A was directly demonstrated by in vivo neutralization of IL-17A. Administration of anti IL-17A antibodies concomitantly to a sub-lethal airway infection with 0.1xLD(50 resulted in a fatal disease.In summary, these data characterize the involvement and underline the protective key role of the IL-17A axis in the lungs from inhalational tularemia.

Are "Market Neutral" Hedge Funds Really Market Neutral?

OpenAIRE

Andrew J. Patton

2009-01-01

Using a variety of different definitions of "neutrality," this study presents significant evidence against the neutrality to market risk of hedge funds in a range of style categories. I generalize standard definitions of "market neutrality," and propose five different neutrality concepts. I suggest statistical tests for each neutrality concept, and apply these tests to a database of monthly returns on 1423 hedge funds from five style categories. For the "market neutral" style, approximately o...

Clostridial Binary Toxins: Iota and C2 Family Portraits

Science.gov (United States)

Stiles, Bradley G.; Wigelsworth, Darran J.; Popoff, Michel R.; Barth, Holger

2011-01-01

There are many pathogenic Clostridium species with diverse virulence factors that include protein toxins. Some of these bacteria, such as C. botulinum, C. difficile, C. perfringens, and C. spiroforme, cause enteric problems in animals as well as humans. These often fatal diseases can partly be attributed to binary protein toxins that follow a classic AB paradigm. Within a targeted cell, all clostridial binary toxins destroy filamentous actin via mono-ADP-ribosylation of globular actin by the A component. However, much less is known about B component binding to cell-surface receptors. These toxins share sequence homology amongst themselves and with those produced by another Gram-positive, spore-forming bacterium also commonly associated with soil and disease: Bacillus anthracis. This review focuses upon the iota and C2 families of clostridial binary toxins and includes: (1) basics of the bacterial source; (2) toxin biochemistry; (3) sophisticated cellular uptake machinery; and (4) host–cell responses following toxin-mediated disruption of the cytoskeleton. In summary, these protein toxins aid diverse enteric species within the genus Clostridium. PMID:22919577

Short Toxin-like Proteins Abound in Cnidaria Genomes

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Michal Linial

2012-11-01

Full Text Available Cnidaria is a rich phylum that includes thousands of marine species. In this study, we focused on Anthozoa and Hydrozoa that are represented by the Nematostella vectensis (Sea anemone and Hydra magnipapillata genomes. We present a method for ranking the toxin-like candidates from complete proteomes of Cnidaria. Toxin-like functions were revealed using ClanTox, a statistical machine-learning predictor trained on ion channel inhibitors from venomous animals. Fundamental features that were emphasized in training ClanTox include cysteines and their spacing along the sequences. Among the 83,000 proteins derived from Cnidaria representatives, we found 170 candidates that fulfill the properties of toxin-like-proteins, the vast majority of which were previously unrecognized as toxins. An additional 394 short proteins exhibit characteristics of toxin-like proteins at a moderate degree of confidence. Remarkably, only 11% of the predicted toxin-like proteins were previously classified as toxins. Based on our prediction methodology and manual annotation, we inferred functions for over 400 of these proteins. Such functions include protease inhibitors, membrane pore formation, ion channel blockers and metal binding proteins. Many of the proteins belong to small families of paralogs. We conclude that the evolutionary expansion of toxin-like proteins in Cnidaria contributes to their fitness in the complex environment of the aquatic ecosystem.

In vitro reconstitution of the Clostridium botulinum type D progenitor toxin.

Science.gov (United States)

Kouguchi, Hirokazu; Watanabe, Toshihiro; Sagane, Yoshimasa; Sunagawa, Hiroyuki; Ohyama, Tohru

2002-01-25

Clostridium botulinum type D strain 4947 produces two different sizes of progenitor toxins (M and L) as intact forms without proteolytic processing. The M toxin is composed of neurotoxin (NT) and nontoxic-nonhemagglutinin (NTNHA), whereas the L toxin is composed of the M toxin and hemagglutinin (HA) subcomponents (HA-70, HA-17, and HA-33). The HA-70 subcomponent and the HA-33/17 complex were isolated from the L toxin to near homogeneity by chromatography in the presence of denaturing agents. We were able to demonstrate, for the first time, in vitro reconstitution of the L toxin formed by mixing purified M toxin, HA-70, and HA-33/17. The properties of reconstituted and native L toxins are indistinguishable with respect to their gel filtration profiles, native-PAGE profiles, hemagglutination activity, binding activity to erythrocytes, and oral toxicity to mice. M toxin, which contained nicked NTNHA prepared by treatment with trypsin, could no longer be reconstituted to the L toxin with HA subcomponents, whereas the L toxin treated with proteases was not degraded into M toxin and HA subcomponents. We conclude that the M toxin forms first by assembly of NT with NTNHA and is subsequently converted to the L toxin by assembly with HA-70 and HA-33/17.

Botulinum toxin in pain treatment.

Science.gov (United States)

Colhado, Orlando Carlos Gomes; Boeing, Marcelo; Ortega, Luciano Bornia

2009-01-01

Botulinum toxin (BTX) is one of the most potent bacterial toxins known and its effectiveness in the treatment of some pain syndromes is well known. However, the efficacy of some of its indications is still in the process of being confirmed. The objective of this study was to review the history, pharmacological properties, and clinical applications of BTX in the treatment of pain of different origins. Botulinum toxin is produced by fermentation of Clostridium botulinum, a Gram-positive, anaerobic bacterium. Commercially, BTX comes in two presentations, types A and B. Botulinum toxin, a neurotoxin with high affinity for cholinergic synapses, blocks the release of acetylcholine by nerve endings without interfering with neuronal conduction of electrical signals or synthesis and storage of acetylcholine. It has been proven that BTX can selectively weaken painful muscles, interrupting the spasm-pain cycle. Several studies have demonstrated the efficacy and safety of BTX-A in the treatment of tension headaches, migraines, chronic lumbar pain, and myofascial pain. Botulinum toxin type A is well tolerated in the treatment of chronic pain disorders in which pharmacotherapy regimens can cause side effects. The reduction in the consumption of analgesics and length of action of 3 to 4 months per dose represent other advantages of its use. However, further studies are necessary to establish the efficacy of BTX-A in chronic pain disorders and its exact mechanism of action, as well as its potential in multifactorial treatments.

Botulinum Toxin for Rhinitis.

Science.gov (United States)

Ozcan, Cengiz; Ismi, Onur

2016-08-01

Rhinitis is a common clinical entity. Besides nasal obstruction, itching, and sneezing, one of the most important symptoms of rhinitis is nasal hypersecretion produced by nasal glands and exudate from the nasal vascular bed. Allergic rhinitis is an IgE-mediated inflammatory reaction of nasal mucosa after exposure to environmental allergens. Idiopathic rhinitis describes rhinitis symptoms that occur after non-allergic, noninfectious irritants. Specific allergen avoidance, topical nasal decongestants, nasal corticosteroids, immunotherapy, and sinonasal surgery are the main treatment options. Because the current treatment modalities are not enough for reducing rhinorrhea in some patients, novel treatment options are required to solve this problem. Botulinum toxin is an exotoxin generated by Clostridium botulinum. It disturbs the signal transmission at the neuromuscular and neuroglandular junction by inhibiting the acetylcholine release from the presynaptic nerve terminal. It has been widely used in neuromuscular, hypersecretory, and autonomic nerve system disorders. There have been a lot of published articles concerning the effect of this toxin on rhinitis symptoms. Based on the results of these reports, intranasal botulinum toxin A administration appears to be a safe and effective treatment method for decreasing rhinitis symptoms in rhinitis patients with a long-lasting effect. Botulinum toxin type A will be a good treatment option for the chronic rhinitis patients who are resistant to other treatment methods.

Diffusion of Botulinum Toxins

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Matthew A. Brodsky

2012-08-01

Full Text Available Background: It is generally agreed that diffusion of botulinum toxin occurs, but the extent of the spread and its clinical importance are disputed. Many factors have been suggested to play a role but which have the most clinical relevance is a subject of much discussion.Methods: This review discusses the variables affecting diffusion, including protein composition and molecular size as well as injection factors (e.g., volume, dose, injection method. It also discusses data on diffusion from comparative studies in animal models and human clinical trials that illustrate differences between the available botulinum toxin products (onabotulinumtoxinA, abobotulinumtoxinA, incobotulinumtoxinA, and rimabotulinumtoxinB.Results: Neither molecular weight nor the presence of complexing proteins appears to affect diffusion; however, injection volume, concentration, and dose all play roles and are modifiable. Both animal and human studies show that botulinum toxin products are not interchangeable, and that some products are associated with greater diffusion and higher rates of diffusion-related adverse events than others.Discussion: Each of the botulinum toxins is a unique pharmacologic entity. A working knowledge of the different serotypes is essential to avoid unwanted diffusion-related adverse events. In addition, clinicians should be aware that the factors influencing diffusion may range from properties intrinsic to the drug to accurate muscle selection as well as dilution, volume, and dose injected.

The role of toxins in Clostridium difficile infection.

Science.gov (United States)

Chandrasekaran, Ramyavardhanee; Lacy, D Borden

2017-11-01

Clostridium difficile is a bacterial pathogen that is the leading cause of nosocomial antibiotic-associated diarrhea and pseudomembranous colitis worldwide. The incidence, severity, mortality and healthcare costs associated with C. difficile infection (CDI) are rising, making C. difficile a major threat to public health. Traditional treatments for CDI involve use of antibiotics such as metronidazole and vancomycin, but disease recurrence occurs in about 30% of patients, highlighting the need for new therapies. The pathogenesis of C. difficile is primarily mediated by the actions of two large clostridial glucosylating toxins, toxin A (TcdA) and toxin B (TcdB). Some strains produce a third toxin, the binary toxin C. difficile transferase, which can also contribute to C. difficile virulence and disease. These toxins act on the colonic epithelium and immune cells and induce a complex cascade of cellular events that result in fluid secretion, inflammation and tissue damage, which are the hallmark features of the disease. In this review, we summarize our current understanding of the structure and mechanism of action of the C. difficile toxins and their role in disease. Published by Oxford University Press on behalf of FEMS 2017.

Temperature Effects Explain Continental Scale Distribution of Cyanobacterial Toxins.

Science.gov (United States)

Mantzouki, Evanthia; Lürling, Miquel; Fastner, Jutta; de Senerpont Domis, Lisette; Wilk-Woźniak, Elżbieta; Koreivienė, Judita; Seelen, Laura; Teurlincx, Sven; Verstijnen, Yvon; Krztoń, Wojciech; Walusiak, Edward; Karosienė, Jūratė; Kasperovičienė, Jūratė; Savadova, Ksenija; Vitonytė, Irma; Cillero-Castro, Carmen; Budzyńska, Agnieszka; Goldyn, Ryszard; Kozak, Anna; Rosińska, Joanna; Szeląg-Wasielewska, Elżbieta; Domek, Piotr; Jakubowska-Krepska, Natalia; Kwasizur, Kinga; Messyasz, Beata; Pełechaty, Aleksandra; Pełechaty, Mariusz; Kokocinski, Mikolaj; García-Murcia, Ana; Real, Monserrat; Romans, Elvira; Noguero-Ribes, Jordi; Duque, David Parreño; Fernández-Morán, Elísabeth; Karakaya, Nusret; Häggqvist, Kerstin; Demir, Nilsun; Beklioğlu, Meryem; Filiz, Nur; Levi, Eti E.; Iskin, Uğur; Bezirci, Gizem; Tavşanoğlu, Ülkü Nihan; Özhan, Koray; Gkelis, Spyros; Panou, Manthos; Fakioglu, Özden; Avagianos, Christos; Kaloudis, Triantafyllos; Çelik, Kemal; Yilmaz, Mete; Marcé, Rafael; Catalán, Nuria; Bravo, Andrea G.; Buck, Moritz; Colom-Montero, William; Mustonen, Kristiina; Pierson, Don; Yang, Yang; Raposeiro, Pedro M.; Gonçalves, Vítor; Antoniou, Maria G.; Tsiarta, Nikoletta; McCarthy, Valerie; Perello, Victor C.; Feldmann, Tõnu; Laas, Alo; Panksep, Kristel; Tuvikene, Lea; Gagala, Ilona; Mankiewicz-Boczek, Joana; Yağcı, Meral Apaydın; Çınar, Şakir; Çapkın, Kadir; Yağcı, Abdulkadir; Cesur, Mehmet; Bilgin, Fuat; Bulut, Cafer; Uysal, Rahmi; Obertegger, Ulrike; Boscaini, Adriano; Flaim, Giovanna; Salmaso, Nico; Cerasino, Leonardo; Richardson, Jessica; Visser, Petra M.; Verspagen, Jolanda M. H.; Karan, Tünay; Soylu, Elif Neyran; Maraşlıoğlu, Faruk; Napiórkowska-Krzebietke, Agnieszka; Ochocka, Agnieszka; Pasztaleniec, Agnieszka; Antão-Geraldes, Ana M.; Vasconcelos, Vitor; Morais, João; Vale, Micaela; Köker, Latife; Akçaalan, Reyhan; Albay, Meriç; Špoljarić Maronić, Dubravka; Stević, Filip; Žuna Pfeiffer, Tanja; Fonvielle, Jeremy; Straile, Dietmar; Rothhaupt, Karl-Otto; Hansson, Lars-Anders; Urrutia-Cordero, Pablo; Bláha, Luděk; Geriš, Rodan; Fránková, Markéta; Koçer, Mehmet Ali Turan; Alp, Mehmet Tahir; Remec-Rekar, Spela; Elersek, Tina; Triantis, Theodoros; Zervou, Sevasti-Kiriaki; Hiskia, Anastasia; Haande, Sigrid; Skjelbred, Birger; Madrecka, Beata; Nemova, Hana; Drastichova, Iveta; Chomova, Lucia; Edwards, Christine; Sevindik, Tuğba Ongun; Tunca, Hatice; Önem, Burçin; Aleksovski, Boris; Krstić, Svetislav; Vucelić, Itana Bokan; Nawrocka, Lidia; Salmi, Pauliina; Machado-Vieira, Danielle; de Oliveira, Alinne Gurjão; Delgado-Martín, Jordi; García, David; Cereijo, Jose Luís; Gomà, Joan; Trapote, Mari Carmen; Vegas-Vilarrúbia, Teresa; Obrador, Biel; Grabowska, Magdalena; Karpowicz, Maciej; Chmura, Damian; Úbeda, Bárbara; Gálvez, José Ángel; Özen, Arda; Christoffersen, Kirsten Seestern; Warming, Trine Perlt; Kobos, Justyna; Mazur-Marzec, Hanna; Pérez-Martínez, Carmen; Ramos-Rodríguez, Eloísa; Arvola, Lauri; Alcaraz-Párraga, Pablo; Toporowska, Magdalena; Pawlik-Skowronska, Barbara; Niedźwiecki, Michał; Pęczuła, Wojciech; Leira, Manel; Hernández, Armand; Moreno-Ostos, Enrique; Blanco, José María; Rodríguez, Valeriano; Montes-Pérez, Jorge Juan; Palomino, Roberto L.; Rodríguez-Pérez, Estela; Carballeira, Rafael; Camacho, Antonio; Picazo, Antonio; Rochera, Carlos; Santamans, Anna C.; Ferriol, Carmen; Romo, Susana; Soria, Juan Miguel; Dunalska, Julita; Sieńska, Justyna; Szymański, Daniel; Kruk, Marek; Kostrzewska-Szlakowska, Iwona; Jasser, Iwona; Žutinić, Petar; Gligora Udovič, Marija; Plenković-Moraj, Anđelka; Frąk, Magdalena; Bańkowska-Sobczak, Agnieszka; Wasilewicz, Michał; Özkan, Korhan; Maliaka, Valentini; Kangro, Kersti; Grossart, Hans-Peter; Paerl, Hans W.; Carey, Cayelan C.; Ibelings, Bas W.

2018-04-13

Insight into how environmental change determines the production and distribution of cyanobacterial toxins is necessary for risk assessment. Management guidelines currently focus on hepatotoxins (microcystins). Increasing attention is given to other classes, such as neurotoxins (e.g., anatoxin-a) and cytotoxins (e.g., cylindrospermopsin) due to their potency. Most studies examine the relationship between individual toxin variants and environmental factors, such as nutrients, temperature and light. In summer 2015, we collected samples across Europe to investigate the effect of nutrient and temperature gradients on the variability of toxin production at a continental scale. Direct and indirect effects of temperature were the main drivers of the spatial distribution in the toxins produced by the cyanobacterial community, the toxin concentrations and toxin quota. Generalized linear models showed that a Toxin Diversity Index (TDI) increased with latitude, while it decreased with water stability. Increases in TDI were explained through a significant increase in toxin variants such as MC-YR, anatoxin and cylindrospermopsin, accompanied by a decreasing presence of MC-LR. While global warming continues, the direct and indirect effects of increased lake temperatures will drive changes in the distribution of cyanobacterial toxins in Europe, potentially promoting selection of a few highly toxic species or strains.

Temperature Effects Explain Continental Scale Distribution of Cyanobacterial Toxins

Directory of Open Access Journals (Sweden)

Evanthia Mantzouki

2018-04-01

Full Text Available Insight into how environmental change determines the production and distribution of cyanobacterial toxins is necessary for risk assessment. Management guidelines currently focus on hepatotoxins (microcystins. Increasing attention is given to other classes, such as neurotoxins (e.g., anatoxin-a and cytotoxins (e.g., cylindrospermopsin due to their potency. Most studies examine the relationship between individual toxin variants and environmental factors, such as nutrients, temperature and light. In summer 2015, we collected samples across Europe to investigate the effect of nutrient and temperature gradients on the variability of toxin production at a continental scale. Direct and indirect effects of temperature were the main drivers of the spatial distribution in the toxins produced by the cyanobacterial community, the toxin concentrations and toxin quota. Generalized linear models showed that a Toxin Diversity Index (TDI increased with latitude, while it decreased with water stability. Increases in TDI were explained through a significant increase in toxin variants such as MC-YR, anatoxin and cylindrospermopsin, accompanied by a decreasing presence of MC-LR. While global warming continues, the direct and indirect effects of increased lake temperatures will drive changes in the distribution of cyanobacterial toxins in Europe, potentially promoting selection of a few highly toxic species or strains.

Lethality of Rendang packaged in multilayer retortable pouch with sterilization process

Science.gov (United States)

Praharasti, A. S.; Kusumaningrum, A.; Frediansyah, A.; Nurhikmat, A.; Khasanah, Y.; Suprapedi

2017-01-01

Retort Pouch had become a choice to preserve foods nowadays, besides the used of the can. Both had their own advantages, and Retort Pouch became more popular for the reason of cheaper and easier to recycle. General Method usually used to estimate the lethality of commercial heat sterilization process. Lethality value wa s used for evaluating the efficacy of the thermal process. This study aimed to find whether different layers of pouch materials affect the lethality value and to find differences lethality in two types of multilayer retort pouch, PET/Aluminum Foil/Nylon/RCPP and PET/Nylon/Modified Aluminum/CPP. The result showed that the different layer arrangement was resulted different Sterilization Value (SV). PET/Nylon/Modified Aluminum/CPP had better heat penetration, implied by the higher value of lethality. PET/Nylon/Modified Aluminum/CPP had the lethality value of 6,24 minutes, whereas the lethality value of PET/Aluminum Foil/Nylon/RCPP was 3,54 minutes.

Genetic fusions of a CFA/I/II/IV MEFA (multiepitope fusion antigen) and a toxoid fusion of heat-stable toxin (STa) and heat-labile toxin (LT) of enterotoxigenic Escherichia coli (ETEC) retain broad anti-CFA and antitoxin antigenicity.

Science.gov (United States)

Ruan, Xiaosai; Sack, David A; Zhang, Weiping

2015-01-01

Immunological heterogeneity has long been the major challenge in developing broadly effective vaccines to protect humans and animals against bacterial and viral infections. Enterotoxigenic Escherichia coli (ETEC) strains, the leading bacterial cause of diarrhea in humans, express at least 23 immunologically different colonization factor antigens (CFAs) and two distinct enterotoxins [heat-labile toxin (LT) and heat-stable toxin type Ib (STa or hSTa)]. ETEC strains expressing any one or two CFAs and either toxin cause diarrhea, therefore vaccines inducing broad immunity against a majority of CFAs, if not all, and both toxins are expected to be effective against ETEC. In this study, we applied the multiepitope fusion antigen (MEFA) strategy to construct ETEC antigens and examined antigens for broad anti-CFA and antitoxin immunogenicity. CFA MEFA CFA/I/II/IV [CVI 2014, 21(2):243-9], which carried epitopes of seven CFAs [CFA/I, CFA/II (CS1, CS2, CS3), CFA/IV (CS4, CS5, CS6)] expressed by the most prevalent and virulent ETEC strains, was genetically fused to LT-STa toxoid fusion monomer 3xSTaA14Q-dmLT or 3xSTaN12S-dmLT [IAI 2014, 82(5):1823-32] for CFA/I/II/IV-STaA14Q-dmLT and CFA/I/II/IV-STaN12S-dmLT MEFAs. Mice intraperitoneally immunized with either CFA/I/II/IV-STa-toxoid-dmLT MEFA developed antibodies specific to seven CFAs and both toxins, at levels equivalent or comparable to those induced from co-administration of the CFA/I/II/IV MEFA and toxoid fusion 3xSTaN12S-dmLT. Moreover, induced antibodies showed in vitro adherence inhibition activities against ETEC or E. coli strains expressing these seven CFAs and neutralization activities against both toxins. These results indicated CFA/I/II/IV-STa-toxoid-dmLT MEFA or CFA/I/II/IV MEFA combined with 3xSTaN12S-dmLT induced broadly protective anti-CFA and antitoxin immunity, and suggested their potential application in broadly effective ETEC vaccine development. This MEFA strategy may be generally used in multivalent

Genetic fusions of a CFA/I/II/IV MEFA (multiepitope fusion antigen and a toxoid fusion of heat-stable toxin (STa and heat-labile toxin (LT of enterotoxigenic Escherichia coli (ETEC retain broad anti-CFA and antitoxin antigenicity.

Directory of Open Access Journals (Sweden)

Xiaosai Ruan

Full Text Available Immunological heterogeneity has long been the major challenge in developing broadly effective vaccines to protect humans and animals against bacterial and viral infections. Enterotoxigenic Escherichia coli (ETEC strains, the leading bacterial cause of diarrhea in humans, express at least 23 immunologically different colonization factor antigens (CFAs and two distinct enterotoxins [heat-labile toxin (LT and heat-stable toxin type Ib (STa or hSTa]. ETEC strains expressing any one or two CFAs and either toxin cause diarrhea, therefore vaccines inducing broad immunity against a majority of CFAs, if not all, and both toxins are expected to be effective against ETEC. In this study, we applied the multiepitope fusion antigen (MEFA strategy to construct ETEC antigens and examined antigens for broad anti-CFA and antitoxin immunogenicity. CFA MEFA CFA/I/II/IV [CVI 2014, 21(2:243-9], which carried epitopes of seven CFAs [CFA/I, CFA/II (CS1, CS2, CS3, CFA/IV (CS4, CS5, CS6] expressed by the most prevalent and virulent ETEC strains, was genetically fused to LT-STa toxoid fusion monomer 3xSTaA14Q-dmLT or 3xSTaN12S-dmLT [IAI 2014, 82(5:1823-32] for CFA/I/II/IV-STaA14Q-dmLT and CFA/I/II/IV-STaN12S-dmLT MEFAs. Mice intraperitoneally immunized with either CFA/I/II/IV-STa-toxoid-dmLT MEFA developed antibodies specific to seven CFAs and both toxins, at levels equivalent or comparable to those induced from co-administration of the CFA/I/II/IV MEFA and toxoid fusion 3xSTaN12S-dmLT. Moreover, induced antibodies showed in vitro adherence inhibition activities against ETEC or E. coli strains expressing these seven CFAs and neutralization activities against both toxins. These results indicated CFA/I/II/IV-STa-toxoid-dmLT MEFA or CFA/I/II/IV MEFA combined with 3xSTaN12S-dmLT induced broadly protective anti-CFA and antitoxin immunity, and suggested their potential application in broadly effective ETEC vaccine development. This MEFA strategy may be generally used in

Neutralization of bitis parviocula (Ethiopian mountain adder venom by the south african institute of medical research (SAIMR antivenom

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Elda E. Sánchez

2011-08-01

Full Text Available BACKGROUND: The Ethiopian mountain adder (Bitis parviocula is a viperid known only from a few locations in southwestern Ethiopia. METHODS: a total of 30 µg of B. arietans and B. parviocula venoms were run on a 10-20% Tricine gel. To assay lethality dose fifty (LD50, five groups of eight mice for each venom were used. Hemorrhagic activity for crude venom was tested. Fibrinogenolytic activity of crude venom was measured using (2.5 mg/mL of fibrinogen solution and (0.03 mg/mL of crude venom. Gelatinase activity of the venom was tested on a Kodak X-OMAT TM film. Crude venoms of B. parviocula and B. arietans were tested for their abilities to affect clotting time, clotting rate and platelet function on whole human blood. RESULTS: The (SAIMR antivenom was confirmed in this study to neutralize the lethal activity of venom from Bitis parviocula. The ED50s of SAIMR antivenom on B. parviocula and B. arietans neutralized half of 18.2 and 66.7 mg of venom, respectively. The hemorrhagic activities (MHDs of B. parviocula and B. arietans were 0.88 and 1.7 µg, respectively. Bitis arietans and B. parviocula venoms degradated α and β chains at different times. The γ chains remained unaffected. Bitis parviocula venom did not exhibit gelatinase activity, while B. arietans had a MGD of 6.9 µg. At 3 mg/mL, the crude venoms of B. parviocula and B. arietans did not significantly affect clotting time or clotting rate. CONCLUSIONS: The SAIMR antivenom is very effective in neutralizing the venom of B. parviocula and should be considered in treating envenomations by these snakes.

[Bladder tumor lethality. Results in the autonomous community of Rioja between 1975-1991].

Science.gov (United States)

Fernández Fernández, A; Gil Fabra, J; Fernández Ruíz, M; Angulo Castellanos, M G; Blanco Martín, E; Otero Mauricio, G

1998-01-01

Between 1975-1991, a total of 557 cases of bladder carcinoma were identified in the Autonomous Community of La Rioja (CAR) which were followed up to December 1994. The overall lethality was 21.9%. 492 cases with 22.35% lethality were identified in males. In females, however, there was 65 cases with 18.46% lethality. The comparison of males and females lethality resulted in p = 0.525. Lethality between cases diagnosed within each 5-year period analyzed is: 1975-1981: 177 cases, lethality 23.72%. 1982-1986: 168 cases, lethality 30.95%. 1987-1991: 212 cases, lethality 13.20%. Between the first and the second 5-year periods, p = 0.132; between the first and third 5-year periods p = 0.007 and between the second and third 5-year periods p CAR for a 22.35% lethality. Lethality is higher in males that in females but the difference is not statistically significant. In the last 5-year period assessed, 1987-1991, a reduction of lethality from bladder neoplasms has been documented.

Self-medication as adaptive plasticity: increased ingestion of plant toxins by parasitized caterpillars.

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Michael S Singer

Full Text Available Self-medication is a specific therapeutic behavioral change in response to disease or parasitism. The empirical literature on self-medication has so far focused entirely on identifying cases of self-medication in which particular behaviors are linked to therapeutic outcomes. In this study, we frame self-medication in the broader realm of adaptive plasticity, which provides several testable predictions for verifying self-medication and advancing its conceptual significance. First, self-medication behavior should improve the fitness of animals infected by parasites or pathogens. Second, self-medication behavior in the absence of infection should decrease fitness. Third, infection should induce self-medication behavior. The few rigorous studies of self-medication in non-human animals have not used this theoretical framework and thus have not tested fitness costs of self-medication in the absence of disease or parasitism. Here we use manipulative experiments to test these predictions with the foraging behavior of woolly bear caterpillars (Grammia incorrupta; Lepidoptera: Arctiidae in response to their lethal endoparasites (tachinid flies. Our experiments show that the ingestion of plant toxins called pyrrolizidine alkaloids improves the survival of parasitized caterpillars by conferring resistance against tachinid flies. Consistent with theoretical prediction, excessive ingestion of these toxins reduces the survival of unparasitized caterpillars. Parasitized caterpillars are more likely than unparasitized caterpillars to specifically ingest large amounts of pyrrolizidine alkaloids. This case challenges the conventional view that self-medication behavior is restricted to animals with advanced cognitive abilities, such as primates, and empowers the science of self-medication by placing it in the domain of adaptive plasticity theory.

Diphtheria toxin translocation across cellular membranes is regulated by sphingolipids

International Nuclear Information System (INIS)

Spilsberg, Bjorn; Hanada, Kentaro; Sandvig, Kirsten

2005-01-01

Diphtheria toxin is translocated across cellular membranes when receptor-bound toxin is exposed to low pH. To study the role of sphingolipids for toxin translocation, both a mutant cell line lacking the first enzyme in de novo sphingolipid synthesis, serine palmitoyltransferase, and a specific inhibitor of the same enzyme, myriocin, were used. The serine palmitoyltransferase-deficient cell line (LY-B) was found to be 10-15 times more sensitive to diphtheria toxin than the genetically complemented cell line (LY-B/cLCB1) and the wild-type cell line (CHO-K1), both when toxin translocation directly across the plasma membrane was induced by exposing cells with surface-bound toxin to low pH, and when the toxin followed its normal route via acidified endosomes into the cytosol. Toxin binding was similar in these three cell lines. Furthermore, inhibition of serine palmitoyltransferase activity by addition of myriocin sensitized the two control cell lines (LY-B/cLCB1 and CHO-K1) to diphtheria toxin, whereas, as expected, no effect was observed in cells lacking serine palmitoyltransferase (LY-B). In conclusion, diphtheria toxin translocation is facilitated by depletion of membrane sphingolipids

Bacterial toxins as pathogen weapons against phagocytes

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Ana edo Vale

2016-02-01

Full Text Available Bacterial toxins are virulence factors that manipulate host cell functions and take over the control of vital processes of living organisms to favour microbial infection. Some toxins directly target innate immune cells, thereby annihilating a major branch of the host immune response. In this review we will focus on bacterial toxins that act from the extracellular milieu and hinder the function of macrophages and neutrophils. In particular, we will concentrate on toxins from Gram-positive and Gram-negative bacteria that manipulate cell signalling or induce cell death by either imposing direct damage to the host cells cytoplasmic membrane or enzymatically modifying key eukaryotic targets. Outcomes regarding pathogen dissemination, host damage and disease progression will be discussed.

Expression of Shiga toxin 2e glycosphingolipid receptors of primary porcine brain endothelial cells and toxin-mediated breakdown of the blood-brain barrier.

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Meisen, Iris; Rosenbrück, Regina; Galla, Hans-Joachim; Hüwel, Sabine; Kouzel, Ivan U; Mormann, Michael; Karch, Helge; Müthing, Johannes

2013-06-01

Shiga toxin (Stx) 2e, released by certain Stx-producing Escherichia coli, is presently the best characterized virulence factor responsible for pig edema disease, which is characterized by hemorrhagic lesions, neurological disorders and often fatal outcomes. Although Stx2e-mediated brain vascular injury is the key event in development of neurologic signs, the glycosphingolipid (GSL) receptors of Stx2e and toxin-mediated impairment of pig brain endothelial cells have not been investigated so far. Here, we report on the detailed structural characterization of Stx2e receptors globotriaosylceramide (Gb3Cer) and globotetraosylceramide (Gb4Cer), which make up the major neutral GSLs in primary porcine brain capillary endothelial cells (PBCECs). Various Gb3Cer and Gb4Cer lipoforms harboring sphingenine (d18:1) or sphinganine (d18:0) and mostly a long-chain fatty acid (C20-C24) were detected. A notable batch-to-batch heterogeneity of primary endothelial cells was observed regarding the extent of ceramide hydroxylation of Gb3Cer or Gb4Cer species. Gb3Cer, Gb4Cer and sphingomyelin preferentially distribute to detergent-resistant membrane fractions and can be considered lipid raft markers in PBCECs. Moreover, we employed an in vitro model of the blood-brain barrier (BBB), which exhibited strong cytotoxic effects of Stx2e on the endothelial monolayer and a rapid collapse of the BBB. These data strongly suggest the involvement of Stx2e in cerebral vascular damage with resultant neurological disturbance characteristic of edema disease.

Prolonged protection against Intranasal challenge with influenza virus following systemic immunization or combinations of mucosal and systemic immunizations with a heat-labile toxin mutant.

Science.gov (United States)

Zhou, Fengmin; Goodsell, Amanda; Uematsu, Yasushi; Vajdy, Michael

2009-04-01

Seasonal influenza virus infections cause considerable morbidity and mortality in the world, and there is a serious threat of a pandemic influenza with the potential to cause millions of deaths. Therefore, practical influenza vaccines and vaccination strategies that can confer protection against intranasal infection with influenza viruses are needed. In this study, we demonstrate that using LTK63, a nontoxic mutant of the heat-labile toxin from Escherichia coli, as an adjuvant for both mucosal and systemic immunizations, systemic (intramuscular) immunization or combinations of mucosal (intranasal) and intramuscular immunizations protected mice against intranasal challenge with a lethal dose of live influenza virus at 3.5 months after the second immunization.

Generation, characterization and epitope mapping of two neutralizing and protective human recombinant antibodies against influenza A H5N1 viruses.

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Lina Sun

Full Text Available BACKGROUND: The development of new therapeutic targets and strategies to control highly pathogenic avian influenza (HPAI H5N1 virus infection in humans is urgently needed. Broadly cross-neutralizing recombinant human antibodies obtained from the survivors of H5N1 avian influenza provide an important role in immunotherapy for human H5N1 virus infection and definition of the critical epitopes for vaccine development. METHODOLOGY/PRINCIPAL FINDINGS: We have characterized two recombinant baculovirus-expressed human antibodies (rhAbs, AVFluIgG01 and AVFluIgG03, generated by screening a Fab antibody phage library derived from a patient recovered from infection with a highly pathogenic avian influenza A H5N1 clade 2.3 virus. AVFluIgG01 cross-neutralized the most of clade 0, clade 1, and clade 2 viruses tested, in contrast, AVFluIgG03 only neutralized clade 2 viruses. Passive immunization of mice with either AVFluIgG01 or AVFluIgG03 antibody resulted in protection from a lethal H5N1 clade 2.3 virus infection. Furthermore, through epitope mapping, we identify two distinct epitopes on H5 HA molecule recognized by these rhAbs and demonstrate their potential to protect against a lethal H5N1 virus infection in a mouse model. CONCLUSIONS/SIGNIFICANCE: Importantly, localization of the epitopes recognized by these two neutralizing and protective antibodies has provided, for the first time, insight into the human antibody responses to H5N1 viruses which contribute to the H5 immunity in the recovered patient. These results highlight the potential of a rhAbs treatment strategy for human H5N1 virus infection and provide new insight for the development of effective H5N1 pandemic vaccines.

Testing the "toxin hypothesis of allergy": Mast cells, IgE, and innate and acquired immune responses to venoms*

Science.gov (United States)

Tsai, Mindy; Starkl, Philipp; Marichal, Thomas; Galli, Stephen J.

2015-01-01

Summary Work in mice indicates that innate functions of mast cells, particularly degradation of venom toxins by mast cell-derived proteases, can enhance resistance to certain arthropod or reptile venoms. Recent reports indicate that acquired Th2 immune responses associated with the production of IgE antibodies, induced by Russell’s viper venom or honeybee venom, or by a component of honeybee venom, bee venom phospholipase 2 (bvPLA2), can increase the resistance of mice to challenge with potentially lethal doses of either of the venoms or bvPLA2. These findings support the conclusion that, in contrast to the detrimental effects associated with allergic Th2 immune responses, mast cells and IgE-dependent immune responses to venoms can contribute to innate and adaptive resistance to venom-induced pathology and mortality. PMID:26210895

Humanitarian Algorithms : A Codified Key Safety Switch Protocol for Lethal Autonomy

OpenAIRE

Nyagudi, Nyagudi Musandu

2014-01-01

With the deployment of lethal autonomous weapons, there is the requirement that any such platform complies with the precepts of International Humanitarian Law. Humanitarian Algorithms[9: p. 9] ensure that lethal autonomous weapon systems perform military/security operations, within the confines of International Humanitarian Law. Unlike other existing techniques of regulating lethal autonomy this scheme advocates for an approach that enables Machine Learning. Lethal autonomous weapons must be ...

Dynamics of plc gene transcription and α-toxin production during growth of Clostridium perfringens strains with contrasting α-toxin production

DEFF Research Database (Denmark)

Abildgaard, Lone; Schramm, Andreas; Rudi, Knut

2009-01-01

The aim of the present study was to investigate transcription dynamics of the α-toxin-encoding plc gene relative to two housekeeping genes (gyrA and rplL) in batch cultures of three Clostridium perfringens strains with low, intermediate, and high levels of α-toxin production, respectively. The plc...... transcript level was always low in the low α-toxin producing strain. For the two other strains, plc transcription showed an inducible pattern and reached a maximum level in the late exponential growth phase. The transcription levels were however inversely correlated to α-toxin production for the two strains....... We propose that this discrepancy is due to differences in plc translation rates between the strains and that strain-specific translational rates therefore must be determined before α-toxin production can be extrapolated from transcript levels in C. perfringens....

Risk Assessment of Shellfish Toxins

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Rex Munday

2013-11-01

Full Text Available Complex secondary metabolites, some of which are highly toxic to mammals, are produced by many marine organisms. Some of these organisms are important food sources for marine animals and, when ingested, the toxins that they produce may be absorbed and stored in the tissues of the predators, which then become toxic to animals higher up the food chain. This is a particular problem with shellfish, and many cases of poisoning are reported in shellfish consumers each year. At present, there is no practicable means of preventing uptake of the toxins by shellfish or of removing them after harvesting. Assessment of the risk posed by such toxins is therefore required in order to determine levels that are unlikely to cause adverse effects in humans and to permit the establishment of regulatory limits in shellfish for human consumption. In the present review, the basic principles of risk assessment are described, and the progress made toward robust risk assessment of seafood toxins is discussed. While good progress has been made, it is clear that further toxicological studies are required before this goal is fully achieved.

Non-Lethal Weapons Program

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Sheets Frequently Asked Questions Non-Lethal Weapons FAQs Active Denial System FAQs Human Electro -Muscular Incapacitation FAQs Related Links Business Opportunities Contact JNLWD Congressional Engagement , Wednesday, Sept 20, 2017. The Active Denial System, blunt-impact munitions, dazzling lasers, LRAD 100X

Induction of Shiga Toxin-Encoding Prophage by Abiotic Environmental Stress in Food.

Science.gov (United States)

Fang, Yuan; Mercer, Ryan G; McMullen, Lynn M; Gänzle, Michael G

2017-10-01

The prophage-encoded Shiga toxin is a major virulence factor in Stx-producing Escherichia coli (STEC). Toxin production and phage production are linked and occur after induction of the RecA-dependent SOS response. However, food-related stress and Stx-prophage induction have not been studied at the single-cell level. This study investigated the effects of abiotic environmental stress on stx expression by single-cell quantification of gene expression in STEC O104:H4 Δ stx2 :: gfp :: amp r In addition, the effect of stress on production of phage particles was determined. The lethality of stressors, including heat, HCl, lactic acid, hydrogen peroxide, and high hydrostatic pressure, was selected to reduce cell counts by 1 to 2 log CFU/ml. The integrity of the bacterial membrane after exposure to stress was measured by propidium iodide (PI). The fluorescent signals of green fluorescent protein (GFP) and PI were quantified by flow cytometry. The mechanism of prophage induction by stress was evaluated by relative gene expression of recA and cell morphology. Acid (pH stress were additionally assessed. H 2 O 2 and mitomycin C induced expression of the prophage and activated a SOS response. In contrast, HCl and lactic acid induced the Stx-prophage but not the SOS response. The lifestyle of STEC exposes the organism to intestinal and extraintestinal environments that impose oxidative and acid stress. A more thorough understanding of the influence of food processing-related stressors on Stx-prophage expression thus facilitates control of STEC in food systems by minimizing prophage induction during food production and storage. Copyright © 2017 American Society for Microbiology.

Toxin gene determination and evolution in scorpaenoid fish.

Science.gov (United States)

Chuang, Po-Shun; Shiao, Jen-Chieh

2014-09-01

In this study, we determine the toxin genes from both cDNA and genomic DNA of four scorpaenoid fish and reconstruct their evolutionary relationship. The deduced protein sequences of the two toxin subunits in Sebastapistes strongia, Scorpaenopsis oxycephala, and Sebastiscus marmoratus are about 700 amino acid, similar to the sizes of the stonefish (Synanceia horrida, and Synanceia verrucosa) and lionfish (Pterois antennata and Pterois volitans) toxins previously published. The intron positions are highly conserved among these species, which indicate the applicability of gene finding by using genomic DNA template. The phylogenetic analysis shows that the two toxin subunits were duplicated prior to the speciation of Scorpaenoidei. The precedence of the gene duplication over speciation indicates that the toxin genes may be common to the whole family of Scorpaeniform. Furthermore, one additional toxin gene has been determined in the genomic DNA of Dendrochirus zebra. The phylogenetic analysis suggests that an additional gene duplication occurred before the speciation of the lionfish (Pteroinae) and a pseudogene may be generally present in the lineage of lionfish. Copyright © 2014 Elsevier Ltd. All rights reserved.

Interactions of Kid-Kis toxin-antitoxin complexes with the parD operator-promotor region of plasmid R1 are piloted by the Kis antitoxin and tuned by the stoichiometry of Kid-Kis oligomers

NARCIS (Netherlands)

Monti, M.C.; Hernandez-Arriaga, A.M.; Kamphuis, M.B.; Lopez-Villarejo, J.; Heck, A.J.R.; Boelens, R.; Diaz-Orejas, R.; van den Heuvel, R.H.H.

2007-01-01

The parD operon of Escherichia coli plasmid R1 encodes a toxin–antitoxin system, which is involved in plasmid stabilization. The toxin Kid inhibits cell growth by RNA degradation and its action is neutralized by the formation of a tight complex with the antitoxin Kis. A fascinating but poorly

Discovery of novel bacterial toxins by genomics and computational biology.

Science.gov (United States)

Doxey, Andrew C; Mansfield, Michael J; Montecucco, Cesare

2018-06-01

Hundreds and hundreds of bacterial protein toxins are presently known. Traditionally, toxin identification begins with pathological studies of bacterial infectious disease. Following identification and cultivation of a bacterial pathogen, the protein toxin is purified from the culture medium and its pathogenic activity is studied using the methods of biochemistry and structural biology, cell biology, tissue and organ biology, and appropriate animal models, supplemented by bioimaging techniques. The ongoing and explosive development of high-throughput DNA sequencing and bioinformatic approaches have set in motion a revolution in many fields of biology, including microbiology. One consequence is that genes encoding novel bacterial toxins can be identified by bioinformatic and computational methods based on previous knowledge accumulated from studies of the biology and pathology of thousands of known bacterial protein toxins. Starting from the paradigmatic cases of diphtheria toxin, tetanus and botulinum neurotoxins, this review discusses traditional experimental approaches as well as bioinformatics and genomics-driven approaches that facilitate the discovery of novel bacterial toxins. We discuss recent work on the identification of novel botulinum-like toxins from genera such as Weissella, Chryseobacterium, and Enteroccocus, and the implications of these computationally identified toxins in the field. Finally, we discuss the promise of metagenomics in the discovery of novel toxins and their ecological niches, and present data suggesting the existence of uncharacterized, botulinum-like toxin genes in insect gut metagenomes. Copyright © 2018. Published by Elsevier Ltd.

In-Vivo Neutralization of Botulinum Neurotoxin Serotype E Using Rabbit Polyclonal Antibody Developed against BoNT/E Light Chain.

Science.gov (United States)

Rani, Sarita; Ponmariappan, S; Sharma, Arti; Kamboj, D V; Jain, A K

2017-01-01

Clostridium botulinum is an obligate anaerobic, Gram positive bacterium that secretes extremely toxic substances known as botulinum neurotoxins (BoNTs) that cause serious paralytic illness called botulism. Based upon the serological properties, these neurotoxin have been classified into seven serotypes designated from A to G. Due to extreme toxicity of BoNTs, these neurotoxins have been designated as category A biowarfare agents. There is no commercial neutralizing antibody available for the treatment of botulism. Hence there is an urgent need to develop therapeutic intervention for prevention and cure of botulism within short period. BoNT antiserum injection is still the effective treatment. In the present study, the recombinant light chain of BoNT/E was successfully purified in soluble form. The purified rBoNT/E LC was used for the generation of polyclonal antibody in rabbit. In order to find out the neutralizing capacity of generated antisera, rabbit antiserum was incubated with 20 LD50 of botulinum neurotoxin type E for 1 hour at 37°C and then injected intraperitoneally (IP) into mice. Further in another set of experiments antiserum was administered in different ways that included administration of - antiserum and BoNT/E toxin simultaneously without preincubation, one after another at the same and different time points for its therapeutic ability. To find out cross neutralization capacity, rBoNT/E LC antiserum was pre-incubated with 5 LD50 of BoNT/A, BoNT/B, BoNT/F and then injected (IP) into mice. In all the cases mice were observed continuously for 96 hours. The results clearly indicate that developed polyclonal rabbit antiserum showed serotype specific neutralization of BoNT/E toxin only but not of BoNT/A, BoNT/B and BoNT/F. The developed antibodies will be used for preventive and therapeutic intervention of type 'E' botulism. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

Non-Lethal Weapons: Opportunities for R&D

Science.gov (United States)

2004-12-01

during the Vietnam War. US; emulsifying agents are used in food processing, drilling fluids, cosmetics , pharmaceuticals, heavy- duty cleaners, textile...conducted in a professional manner, with no threat to public safety or the environment. 11 References [1] Fenton , G., (2001). NLW Technology Taxonomy...W.A., Mason, R.L., Collins, K.R., (2000). Non-Lethal Applicants of Slippery Substances. NDIA Non-Lethal Defense IV. [24] Fenton , G., (2000). Overview

77 FR 9888 - Shiga Toxin-Producing Escherichia coli

Science.gov (United States)

2012-02-21

... Toxin-Producing Escherichia coli in Certain Raw Beef Products AGENCY: Food Safety and Inspection Service... toxin-producing Escherichia coli (STEC) serogroups (O26, O45, O103, O111, O121, and O145). This new date..., that are contaminated with Shiga toxin-producing Escherichia coli (STEC) O26, O45, O103, O111, O121...

Cellular Uptake of the Clostridium perfringens Binary Iota-Toxin

Science.gov (United States)

Blöcker, Dagmar; Behlke, Joachim; Aktories, Klaus; Barth, Holger

2001-01-01

The binary iota-toxin is produced by Clostridium perfringens type E strains and consists of two separate proteins, the binding component iota b (98 kDa) and an actin-ADP-ribosylating enzyme component iota a (47 kDa). Iota b binds to the cell surface receptor and mediates the translocation of iota a into the cytosol. Here we studied the cellular uptake of iota-toxin into Vero cells. Bafilomycin A1, but not brefeldin A or nocodazole, inhibited the cytotoxic effects of iota-toxin, indicating that toxin is translocated from an endosomal compartment into the cytoplasm. Acidification (pH ≤ 5.0) of the extracellular medium enabled iota a to directly enter the cytosol in the presence of iota b. Activation by chymotrypsin induced oligomerization of iota b in solution. An average mass of 530 ± 28 kDa for oligomers was determined by analytical ultracentrifugation, indicating heptamer formation. The entry of iota-toxin into polarized CaCo-2 cells was studied by measuring the decrease in transepithelial resistance after toxin treatment. Iota-toxin led to a significant decrease in resistance when it was applied to the basolateral surface of the cells but not following application to the apical surface, indicating a polarized localization of the iota-toxin receptor. PMID:11292715

T-2 toxin Analysis in Poultry and Cattle Feedstuff.

Science.gov (United States)

Gholampour Azizi, Issa; Azarmi, Masumeh; Danesh Pouya, Naser; Rouhi, Samaneh

2014-05-01

T-2 toxin is a mycotoxin that is produced by the Fusarium fungi. Consumption of food and feed contaminated with T-2 toxin causes diseases in humans and animals. In this study T-2 toxin was analyzed in poultry and cattle feedstuff in cities of Mazandaran province (Babol, Sari, Chalus), Northern Iran. In this study, 90 samples were analyzed for T-2 toxin contamination by the ELISA method. Out of 60 concentrate and bagasse samples collected from various cities of Mazandaran province, 11.7% and 3.3% were contaminated with T-2 toxin at concentrations > 25 and 50 µg/kg, respectively. For mixed poultry diets, while 10% of the 30 analyzed samples were contaminated with > 25 µg/kg, none of the tested samples contained T-2 toxin at levels > 50 µg/kg. The results obtained from this study show that poultry and cattle feedstuff can be contaminated with different amounts of T-2 toxin in different conditions and locations. Feedstuff that are contaminated by this toxin cause different diseases in animals; thus, potential transfer of mycotoxins to edible by-products from animals fed mycotoxin-contaminated feeds drives the need to routinely monitor mycotoxins in animal feeds and their components. This is the basis on which effective management of mycotoxins and their effects can be implemented.

Dinophysis Toxins: Causative Organisms, Distribution and Fate in Shellfish

Science.gov (United States)

Reguera, Beatriz; Riobó, Pilar; Rodríguez, Francisco; Díaz, Patricio A.; Pizarro, Gemita; Paz, Beatriz; Franco, José M.; Blanco, Juan

2014-01-01

Several Dinophysis species produce diarrhoetic toxins (okadaic acid and dinophysistoxins) and pectenotoxins, and cause gastointestinal illness, Diarrhetic Shellfish Poisoning (DSP), even at low cell densities (Chile, and Europe. Toxicity and toxin profiles are very variable, more between strains than species. The distribution of DSP events mirrors that of shellfish production areas that have implemented toxin regulations, otherwise misinterpreted as bacterial or viral contamination. Field observations and laboratory experiments have shown that most of the toxins produced by Dinophysis are released into the medium, raising questions about the ecological role of extracelular toxins and their potential uptake by shellfish. Shellfish contamination results from a complex balance between food selection, adsorption, species-specific enzymatic transformations, and allometric processes. Highest risk areas are those combining Dinophysis strains with high cell content of okadaates, aquaculture with predominance of mytilids (good accumulators of toxins), and consumers who frequently include mussels in their diet. Regions including pectenotoxins in their regulated phycotoxins will suffer from much longer harvesting bans and from disloyal competition with production areas where these toxins have been deregulated. PMID:24447996

Lethal synergy involving bicyclomycin: an approach for reviving old antibiotics.

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Malik, Muhammad; Li, Liping; Zhao, Xilin; Kerns, Robert J; Berger, James M; Drlica, Karl

2014-12-01

One way to address the growing problem of antimicrobial resistance is to revive old compounds that may have intrinsic lethal activity that is obscured by protective factors. Bicyclomycin is an old inhibitor of the Rho transcription terminator that by itself shows little rapid lethal activity. However, bicyclomycin participates in bacteriostatic synergy, which raises the possibility that conditions for lethal synergy may exist, perhaps through a suppression of protective factors. Bicyclomycin was combined with bacteriostatic inhibitors of gene expression, and bactericidal activity was measured with several cultured Gram-negative pathogens. When used alone, bicyclomycin failed to rapidly kill growing cultures of Escherichia coli; however, the additional presence of bacteriostatic concentrations of tetracycline, chloramphenicol or rifampicin led to rapid killing. Four other pathogen species, Acinetobacter baumannii, Klebsiella pneumoniae, Salmonella enterica serotype Typhimurium and Shigella dysenteriae, also exhibited enhanced killing when bicyclomycin was combined with tetracycline or rifampicin. This lethal synergy was achieved at low concentrations (slightly above the MIC) for all agents tested in combinations. Follow-up work with E. coli indicated that lethal synergy arose from a blockage of transcription elongation. Moreover, lethal synergy was reduced when bicyclomycin was added 60 min before tetracycline, suggesting that bicyclomycin induces a protective factor. The action of bicyclomycin illustrates the potential present in a largely abandoned antibacterial agent; it exhibits lethal synergy when coadministered with known, bacteriostatic inhibitors of gene expression. The identification of protective factors, which are currently uncharacterized, may reveal new ways to promote the lethal action of some old antibiotics. © The Author 2014. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy. All rights reserved

Isolation of Shiga toxin-producing Escherichia coli harboring variant Shiga toxin genes from seafood

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Sreepriya Prakasan

2018-03-01

Full Text Available Background and Aim: Shiga toxin-producing Escherichia coli (STEC are important pathogens of global significance. STEC are responsible for numerous food-borne outbreaks worldwide and their presence in food is a potential health hazard. The objective of the present study was to determine the incidence of STEC in fresh seafood in Mumbai, India, and to characterize STEC with respect to their virulence determinants. Materials and Methods: A total of 368 E. coli were isolated from 39 fresh seafood samples (18 finfish and 21 shellfish using culture-based methods. The isolates were screened by polymerase chain reaction (PCR for the genes commonly associated with STEC. The variant Shiga toxin genes were confirmed by Southern blotting and hybridization followed by DNA sequencing. Results: One or more Shiga toxins genes were detected in 61 isolates. Of 39 samples analyzed, 10 (25.64% samples harbored STEC. Other virulence genes, namely, eaeA (coding for an intimin and hlyA (hemolysin A were detected in 43 and 15 seafood isolates, respectively. The variant stx1 genes from 6 isolates were sequenced, five of which were found to be stx1d variants, while one sequence varied considerably from known stx1 sequences. Southern hybridization and DNA sequence analysis suggested putative Shiga toxin variant genes (stx2 in at least 3 other isolates. Conclusion: The results of this study showed the occurrence of STEC in seafood harboring one or more Shiga toxin genes. The detection of STEC by PCR may be hampered due to the presence of variant genes such as the stx1d in STEC. This is the first report of stx1d gene in STEC isolated from Indian seafood.

VapC toxins from Mycobacterium tuberculosis are ribonucleases that differentially inhibit growth and are neutralized by cognate VapB antitoxins.

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Bintou Ahmadou Ahidjo

Full Text Available The chromosome of Mycobacterium tuberculosis (Mtb encodes forty seven toxin-antitoxin modules belonging to the VapBC family. The role of these modules in the physiology of Mtb and the function(s served by their expansion are unknown. We investigated ten vapBC modules from Mtb and the single vapBC from M. smegmatis. Of the Mtb vapCs assessed, only Rv0549c, Rv0595c, Rv2549c and Rv2829c were toxic when expressed from a tetracycline-regulated promoter in M. smegmatis. The same genes displayed toxicity when conditionally expressed in Mtb. Toxicity of Rv2549c in M. smegmatis correlated with the level of protein expressed, suggesting that the VapC level must exceed a threshold for toxicity to be observed. In addition, the level of Rv2456 protein induced in M. smegmatis was markedly lower than Rv2549c, which may account for the lack of toxicity of this and other VapCs scored as 'non-toxic'. The growth inhibitory effects of toxic VapCs were neutralized by expression of the cognate VapB as part of a vapBC operon or from a different chromosomal locus, while that of non-cognate antitoxins did not. These results demonstrated a specificity of interaction between VapCs and their cognate VapBs, a finding corroborated by yeast two-hybrid analyses. Deletion of selected vapC or vapBC genes did not affect mycobacterial growth in vitro, but rendered the organisms more susceptible to growth inhibition following toxic VapC expression. However, toxicity of 'non-toxic' VapCs was not unveiled in deletion mutant strains, even when the mutation eliminated the corresponding cognate VapB, presumably due to insufficient levels of VapC protein. Together with the ribonuclease (RNase activity demonstrated for Rv0065 and Rv0617--VapC proteins with similarity to Rv0549c and Rv3320c, respectively--these results suggest that the VapBC family potentially provides an abundant source of RNase activity in Mtb, which may profoundly impact the physiology of the organism.

[Environmental toxins in breast milk].

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Bratlid, Dag

2009-12-17

Breast milk is very important to ensure infants a well-composed and safe diet during the first year of life. However, the quality of breast milk seems to be affected by an increasing amount of environmental toxins (particularly so-called Persistent, Bioaccumulative Toxins [PBTs]). Many concerns have been raised about the negative effects this may have on infant health. The article is a review of literature (mainly review articles) identified through a non-systematic search in PubMed. The concentration of PBTs in breast milk is mainly caused by man's position as the terminal link in the nutritional chain. Many breast-fed infants have a daily intake of such toxins that exceed limits defined for the population in general. Animal studies demonstrate effects on endocrine function and neurotoxicity in the offspring, and a number of human studies seem to point in the same direction. However the "original" optimal composition of breast milk still seems to protect against long-term effects of such toxicity. There is international consensus about the need to monitor breast milk for the presence of PBTs. Such surveillance will be a good indicator of the population's general exposure to these toxins and may also contribute to identifying groups as risk who should not breast-feed their children for a long time.

Botulinum toxin for treatment of glandular hypersecretory disorders.

LENUS (Irish Health Repository)

Laing, T A

2012-02-03

SUMMARY: The use of botulinum toxin to treat disorders of the salivary glands is increasing in popularity in recent years. Recent reports of the use of botulinum toxin in glandular hypersecretion suggest overall favourable results with minimal side-effects. However, few randomised clinical trials means that data are limited with respect to candidate suitability, treatment dosages, frequency and duration of treatment. We report a selection of such cases from our own department managed with botulinum toxin and review the current data on use of the toxin to treat salivary gland disorders such as Frey\\'s syndrome, excessive salivation (sialorrhoea), focal and general hyperhidrosis, excessive lacrimation and chronic rhinitis.

Neutralizing antibodies induced by recombinant virus-like particles of enterovirus 71 genotype C4 inhibit infection at pre- and post-attachment steps.

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Zhiqiang Ku

Full Text Available BACKGROUND: Enterovirus 71 (EV71 is a major causative agent of hand, foot and mouth disease, which has been prevalent in Asia-Pacific regions, causing significant morbidity and mortality in young children. Antibodies elicited by experimental EV71 vaccines could neutralize infection in vitro and passively protect animal models from lethal challenge, indicating that neutralizing antibodies play an essential role in protection. However, how neutralizing antibodies inhibit infection in vitro remains unclear. METHODS/FINDINGS: In the present study, we explored the mechanisms of neutralization by antibodies against EV71 virus-like particles (VLPs. Recombinant VLPs of EV71 genotype C4 were produced in insect cells using baculovirus vectors. Immunization with the VLPs elicited a high-titer, EV71-specific antibody response in mice. Anti-VLP mouse sera potently neutralized EV71 infection in vitro. The neutralizing antibodies in the anti-VLP mouse sera were found to target mainly an extremely conserved epitope (FGEHKQEKDLEYGAC located at the GH loop of the VP1 protein. The neutralizing anti-VLP antisera were able to inhibit virus binding to target cells efficiently. In addition, post-attachment treatment of virus-bound cells with the anti-VLP antisera also neutralized virus infection, although the antibody concentration required was higher than that of the pre-attachment treatment. CONCLUSIONS: Collectively, our findings represent a valuable addition to the understanding of mechanisms of EV71 neutralization and have strong implications for EV71 vaccine development.

Characterization of a Toxin A-Negative, Toxin B-Positive Strain of Clostridium difficile Responsible for a Nosocomial Outbreak of Clostridium difficile-Associated Diarrhea

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Alfa, Michelle J.; Kabani, Amin; Lyerly, David; Moncrief, Scott; Neville, Laurie M.; Al-Barrak, Ali; Harding, Godfrey K. H.; Dyck, Brenda; Olekson, Karen; Embil, John M.

2000-01-01

Clostridium difficile-associated diarrhea (CAD) is a very common nosocomial infection that contributes significantly to patient morbidity and mortality as well as to the cost of hospitalization. Previously, strains of toxin A-negative, toxin B-positive C. difficile were not thought to be associated with clinically significant disease. This study reports the characterization of a toxin A-negative, toxin B-positive strain of C. difficile that was responsible for a recently described nosocomial outbreak of CAD. Analysis of the seven patient isolates from the outbreak by pulsed-field gel electrophoresis indicated that this outbreak was due to transmission of a single strain of C. difficile. Our characterization of this strain (HSC98) has demonstrated that the toxin A gene lacks 1.8 kb from the carboxy repetitive oligopeptide (CROP) region but apparently has no other major deletions from other regions of the toxin A or toxin B gene. The remaining 1.3-kb fragment of the toxin A CROP region from strain HSC98 showed 98% sequence homology with strain 1470, previously reported by M. Weidmann in 1997 (GenBank accession number Y12616), suggesting that HSC98 is toxinotype VIII. The HSC98 strain infecting patients involved in this outbreak produced the full spectrum of clinical illness usually associated with C. difficile-associated disease. This pathogenic spectrum was manifest despite the inability of this strain to alter tight junctions as determined by using in vitro tissue culture testing, which suggested that no functional toxin A was produced by this strain. PMID:10878068

Uptake and bioaccumulation of Cry toxins by an aphidophagous predator

International Nuclear Information System (INIS)

Paula, Débora P.; Andow, David A.

2016-01-01

Uptake of Cry toxins by insect natural enemies has rarely been considered and bioaccumulation has not yet been demonstrated. Uptake can be demonstrated by the continued presence of Cry toxin after exposure has stopped and gut contents eliminated. Bioaccumulation can be demonstrated by showing uptake and that the concentration of Cry toxin in the natural enemy exceeds that in its food. We exposed larvae of the aphidophagous predator, Harmonia axyridis, to Cry1Ac and Cry1F through uniform and constant tritrophic exposure via an aphid, Myzus persicae, and looked for toxin presence in the pupae. We repeated the experiment using only Cry1F and tested newly emerged adults. Both Cry toxins were detected in pupae, and Cry1F was detected in recently emerged, unfed adults. Cry1Ac was present 2.05 times and Cry1F 3.09 times higher in predator pupae than in the aphid prey. Uptake and bioaccumulation in the third trophic level might increase the persistence of Cry toxins in the food web and mediate new exposure routes to natural enemies. - Highlights: • Uptake and bioaccumulation of two Cry toxins by a larval coccinellid was tested. • Uptake was demonstrated by presence of the toxins in pupae and adults. • Bioaccumulation was shown by higher toxin concentration in pupae than prey. • Cry1Ac was present 2.05× and Cry1F 3.09× higher in predator pupae than prey. • This might increase persistence of Cry toxins in food webs with new exposure routes. - Immatures of the predaceous coccinellid Harmonia axyridis can uptake and bioaccumulate Cry toxins delivered via their aphid prey.

[Botulinum toxin: An important complement for facial rejuvenation surgery].

Science.gov (United States)

Le Louarn, C

2017-10-01

The improved understanding of the functional anatomy of the face and of the action of the botulinum toxin A leads us to determine a new injection procedure which consequently decreases the risk of eyebrow and eyelid ptosis and increases the toxin's injection possibilities and efficiencies. With less units of toxin, the technique herein described proposes to be more efficient on more muscles: variable toxin injections concentration adapted to each injected muscle are used. Thanks to a new procedure in the upper face, toxin A injection can be quite close to an endoscopic surgical action. In addition, interesting results are achievable to rejuvenate the lateral canthus with injection on the upper lateral tarsus, to rejuvenate the nose with injection at the alar base, the jawline and the neck region. Lastly, a smoothing effect on the skin (meso botox) is obtained by the anticholinergic action of the toxin A on the dermal receptors. Copyright © 2017. Published by Elsevier Masson SAS.

AB toxins: a paradigm switch from deadly to desirable.

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Odumosu, Oludare; Nicholas, Dequina; Yano, Hiroshi; Langridge, William

2010-07-01

To ensure their survival, a number of bacterial and plant species have evolved a common strategy to capture energy from other biological systems. Being imperfect pathogens, organisms synthesizing multi-subunit AB toxins are responsible for the mortality of millions of people and animals annually. Vaccination against these organisms and their toxins has proved rather ineffective in providing long-term protection from disease. In response to the debilitating effects of AB toxins on epithelial cells of the digestive mucosa, mechanisms underlying toxin immunomodulation of immune responses have become the focus of increasing experimentation. The results of these studies reveal that AB toxins may have a beneficial application as adjuvants for the enhancement of immune protection against infection and autoimmunity. Here, we examine similarities and differences in the structure and function of bacterial and plant AB toxins that underlie their toxicity and their exceptional properties as immunomodulators for stimulating immune responses against infectious disease and for immune suppression of organ-specific autoimmunity.

AB Toxins: A Paradigm Switch from Deadly to Desirable

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Oludare Odumosu

2010-06-01

Full Text Available To ensure their survival, a number of bacterial and plant species have evolved a common strategy to capture energy from other biological systems. Being imperfect pathogens, organisms synthesizing multi-subunit AB toxins are responsible for the mortality of millions of people and animals annually. Vaccination against these organisms and their toxins has proved rather ineffective in providing long-term protection from disease. In response to the debilitating effects of AB toxins on epithelial cells of the digestive mucosa, mechanisms underlying toxin immunomodulation of immune responses have become the focus of increasing experimentation. The results of these studies reveal that AB toxins may have a beneficial application as adjuvants for the enhancement of immune protection against infection and autoimmunity. Here, we examine similarities and differences in the structure and function of bacterial and plant AB toxins that underlie their toxicity and their exceptional properties as immunomodulators for stimulating immune responses against infectious disease and for immune suppression of organ-specific autoimmunity.

Botulinum toxin A for the Treatment of Overactive Bladder.

Science.gov (United States)

Hsieh, Po-Fan; Chiu, Hung-Chieh; Chen, Kuan-Chieh; Chang, Chao-Hsiang; Chou, Eric Chieh-Lung

2016-02-29

The standard treatment for overactive bladder starts with patient education and behavior therapies, followed by antimuscarinic agents. For patients with urgency urinary incontinence refractory to antimuscarinic therapy, currently both American Urological Association (AUA) and European Association of Urology (EAU) guidelines suggested that intravesical injection of botulinum toxin A should be offered. The mechanism of botulinum toxin A includes inhibition of vesicular release of neurotransmitters and the axonal expression of capsaicin and purinergic receptors in the suburothelium, as well as attenuation of central sensitization. Multiple randomized, placebo-controlled trials demonstrated that botulinum toxin A to be an effective treatment for patients with refractory idiopathic or neurogenic detrusor overactivity. The urinary incontinence episodes, maximum cystometric capacity, and maximum detrusor pressure were improved greater by botulinum toxin A compared to placebo. The adverse effects of botulinum toxin A, such as urinary retention and urinary tract infection, were primarily localized to the lower urinary tract. Therefore, botulinum toxin A offers an effective treatment option for patients with refractory overactive bladder.

Toxins and antimicrobial peptides: interactions with membranes

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Schlamadinger, Diana E.; Gable, Jonathan E.; Kim, Judy E.

2009-08-01

The innate immunity to pathogenic invasion of organisms in the plant and animal kingdoms relies upon cationic antimicrobial peptides (AMPs) as the first line of defense. In addition to these natural peptide antibiotics, similar cationic peptides, such as the bee venom toxin melittin, act as nonspecific toxins. Molecular details of AMP and peptide toxin action are not known, but the universal function of these peptides to disrupt cell membranes of pathogenic bacteria (AMPs) or a diverse set of eukaryotes and prokaryotes (melittin) is widely accepted. Here, we have utilized spectroscopic techniques to elucidate peptide-membrane interactions of alpha-helical human and mouse AMPs of the cathelicidin family as well as the peptide toxin melittin. The activity of these natural peptides and their engineered analogs was studied on eukaryotic and prokaryotic membrane mimics consisting of resistant pathogens.

Botulinum toxin type a for chronic migraine.

Science.gov (United States)

Ashkenazi, Avi

2010-03-01

Chronic migraine (CM) is the leading cause of chronic daily headache, a common and debilitating headache syndrome. The management of CM patients is challenging, with only limited benefit from available oral preventive medications. Botulinum neurotoxin (BoNT) has been used extensively to treat disorders associated with increased muscle tone. More recent scientific data support an analgesic effect of the toxin. The pharmacokinetic and pharmacodynamic profiles of BoNT make it an appealing candidate for migraine prevention. Results from older clinical trials on the efficacy of the toxin in CM were inconclusive. However, recent trials using more stringent inclusion criteria have shown positive results, supporting the use of the toxin in some patients with this disorder. This review summarizes the scientific data on the analgesic properties of BoNT, as well as the clinical data on the efficacy of the toxin in treating CM.

Integrated microfluidic technology for sub-lethal and behavioral marine ecotoxicity biotests

Science.gov (United States)

Huang, Yushi; Reyes Aldasoro, Constantino Carlos; Persoone, Guido; Wlodkowic, Donald

2015-06-01

Changes in behavioral traits exhibited by small aquatic invertebrates are increasingly postulated as ethically acceptable and more sensitive endpoints for detection of water-born ecotoxicity than conventional mortality assays. Despite importance of such behavioral biotests, their implementation is profoundly limited by the lack of appropriate biocompatible automation, integrated optoelectronic sensors, and the associated electronics and analysis algorithms. This work outlines development of a proof-of-concept miniaturized Lab-on-a-Chip (LOC) platform for rapid water toxicity tests based on changes in swimming patterns exhibited by Artemia franciscana (Artoxkit M™) nauplii. In contrast to conventionally performed end-point analysis based on counting numbers of dead/immobile specimens we performed a time-resolved video data analysis to dynamically assess impact of a reference toxicant on swimming pattern of A. franciscana. Our system design combined: (i) innovative microfluidic device keeping free swimming Artemia sp. nauplii under continuous microperfusion as a mean of toxin delivery; (ii) mechatronic interface for user-friendly fluidic actuation of the chip; and (iii) miniaturized video acquisition for movement analysis of test specimens. The system was capable of performing fully programmable time-lapse and video-microscopy of multiple samples for rapid ecotoxicity analysis. It enabled development of a user-friendly and inexpensive test protocol to dynamically detect sub-lethal behavioral end-points such as changes in speed of movement or distance traveled by each animal.

Stealth and mimicry by deadly bacterial toxins

DEFF Research Database (Denmark)

Yates, S.P.; Jørgensen, Rene; Andersen, Gregers Rom

2006-01-01

Diphtheria toxin and exotoxin A are well-characterized members of the ADP-ribosyltransferase toxin family that serve as virulence factors in the pathogenic bacteria, Corynebacterium diphtheriae and Pseudomonas aeruginosa.  New high-resolution structural data of the Michaelis complex...

Acid Sphingomyelinase Promotes Cellular Internalization of Clostridium perfringens Iota-Toxin.

Science.gov (United States)

Nagahama, Masahiro; Takehara, Masaya; Miyamoto, Kazuaki; Ishidoh, Kazumi; Kobayashi, Keiko

2018-05-20

Clostridium perfringens iota-toxin is a binary actin-ADP-ribosylating toxin composed of the enzymatic component Ia and receptor binding component Ib. Ib binds to a cell surface receptor, forms Ib oligomer in lipid rafts, and associates with Ia. The Ia-Ib complex then internalizes by endocytosis. Here, we showed that acid sphingomyelinase (ASMase) facilitates the cellular uptake of iota-toxin. Inhibitions of ASMase and lysosomal exocytosis by respective blockers depressed cell rounding induced by iota-toxin. The cytotoxicity of the toxin increased in the presence of Ca 2+ in extracellular fluids. Ib entered target cells in the presence but not the absence of Ca 2+ . Ib induced the extracellular release of ASMase in the presence of Ca 2+ . ASMase siRNA prevented the cell rounding induced by iota-toxin. Furthermore, treatment of the cells with Ib resulted in the production of ceramide in cytoplasmic vesicles. These observations showed that ASMase promotes the internalization of iota-toxin into target cells.

Influence Analysis of Shell Material and Charge on Shrapnel Lethal Power

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Wang Lin

2015-01-01

Full Text Available To compare the shrapnel lethal power with different shell material and charge, LS-DYNA was used to numerically simulate four kinds of shrapnel lethal power. The shell material was 58SiMn, 50SiMnVB or 40Cr, whereas the charge was RL-F. And the shell material was 58SiMn, whereas the charge was TNT. The shell rupture process and lethal power test were analyzed. The results show that, the lethal power of RL-F charge increase by 25%, 45%, 14% compared with the TNT charge, whereas the shell material was 58SiMn, 50SiMnVB, 40Cr. And then the guarantee range and lethal power can be improved by using the high explosive and changing shell material, whereas the projectile shape coefficient is invariable.

Solid-phase synthesis of polyamine toxin analogues

DEFF Research Database (Denmark)

Kromann, Hasse; Krikstolaityte, Sonata; Andersen, Anne J

2002-01-01

The wasp toxin philanthotoxin-433 (PhTX-433) is a nonselective and noncompetitive antagonist of ionotropic receptors, such as ionotropic glutamate receptors and nicotinic acetylcholine receptors. Polyamine toxins are extensively used for the characterization of subtypes of ionotropic glutamate re...

Cnidarian Toxins Acting on Voltage-Gated Ion Channels

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Robert M. Greenberg

2006-04-01

Full Text Available Abstract: Voltage-gated ion channels generate electrical activity in excitable cells. As such, they are essential components of neuromuscular and neuronal systems, and are targeted by toxins from a wide variety of phyla, including the cnidarians. Here, we review cnidarian toxins known to target voltage-gated ion channels, the specific channel types targeted, and, where known, the sites of action of cnidarian toxins on different channels.

Recombinant thrombomodulin protects mice against histone-induced lethal thromboembolism.

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Mayumi Nakahara

Full Text Available INTRODUCTION: Recent studies have shown that histones, the chief protein component of chromatin, are released into the extracellular space during sepsis, trauma, and ischemia-reperfusion injury, and act as major mediators of the death of an organism. This study was designed to elucidate the cellular and molecular basis of histone-induced lethality and to assess the protective effects of recombinant thrombomodulin (rTM. rTM has been approved for the treatment of disseminated intravascular coagulation (DIC in Japan, and is currently undergoing a phase III clinical trial in the United States. METHODS: Histone H3 levels in plasma of healthy volunteers and patients with sepsis and DIC were measured using enzyme-linked immunosorbent assay. Male C57BL/6 mice were injected intravenously with purified histones, and pathological examinations were performed. The protective effects of rTM against histone toxicity were analyzed both in vitro and in mice. RESULTS: Histone H3 was not detectable in plasma of healthy volunteers, but significant levels were observed in patients with sepsis and DIC. These levels were higher in non-survivors than in survivors. Extracellular histones triggered platelet aggregation, leading to thrombotic occlusion of pulmonary capillaries and subsequent right-sided heart failure in mice. These mice displayed symptoms of DIC, including thrombocytopenia, prolonged prothrombin time, decreased fibrinogen, fibrin deposition in capillaries, and bleeding. Platelet depletion protected mice from histone-induced death in the first 30 minutes, suggesting that vessel occlusion by platelet-rich thrombi might be responsible for death during the early phase. Furthermore, rTM bound to extracellular histones, suppressed histone-induced platelet aggregation, thrombotic occlusion of pulmonary capillaries, and dilatation of the right ventricle, and rescued mice from lethal thromboembolism. CONCLUSIONS: Extracellular histones cause massive

Inhibitory action of chlorophyllin of autosome recessive lethals induced by irradiation

International Nuclear Information System (INIS)

Salceda, V.M.; Pimentel, P.A.E.; Cruces, M.P.

2006-01-01

The chlorophyllin is a sodium salt of the chlorophyll that has a strong protective action of the damage induced by different agents so much physical as chemical. In Drosophila there is reported this effect in somatic cells. In contrast, in germinal cells using tests with the sexual chromosomes has not been found such inhibitory action. For this reason, in this occasion we will refer to the effect of the lethality induced in autosome chromosomes, in particular to the chromosome II of this species. For such effect groups of males of the line Canton-S its were pre-treated for 24h with or without 69 mm of CCS and later on treaties with or without 40 Gy of gamma irradiation. The males were then subjected to the technical Cy L / Pm for the detection of recessive lethals. In the third generation the respective counts of the descendant of each one of them to determine the corresponding categories for each extracted chromosome were made. To be mendelian crosses it is expected for a normal chromosome a proportion 2:1 of individuals with genotype Cy L / +: +/+. The absence of individuals +/+ it is indicative of a lethal gene, until 10% of these individuals of each male's total descendant, it is considered that is carrying of a semi lethal gene. The sum of lethal and semi lethals constitutes the category detrimental. The obtained results indicated that the pre-treatment with CCS reduces in a significant way the frequency of induced lethals by 40 Gy of gamma rays. The fact that an effect inhibitor has not been observed in the test of recessive lethal bound to the sex obtained previously, it contrasts with the effect observed in the chromosome II, results of this study and with the one observed in the chromosome III in somatic cells. The above-mentioned shows a differential action of the CCS between sexual chromosomes and autosomal before the effect of the gamma radiation. At the moment we don't have an explanation to these evidences. To evaluate the action of the chlorophyllin

Lethal and Sub-lethal Effects of Four Insecticides on the Aphidophagous Coccinellid Adalia bipunctata (Coleoptera: Coccinellidae).

Science.gov (United States)

Depalo, Laura; Lanzoni, Alberto; Masetti, Antonio; Pasqualini, Edison; Burgio, Giovanni

2017-12-05

Conventional insecticide assays, which measure the effects of insecticide exposure on short-term mortality, overlook important traits, including persistence of toxicity or sub-lethal effects. Therefore, such approaches are especially inadequate for prediction of the overall impact of insecticides on beneficial arthropods. In this study, the side effects of four modern insecticides (chlorantraniliprole, emamectin benzoate, spinosad, and spirotetramat) on Adalia bipunctata (L.) (Coleoptera: Coccinellidae) were evaluated under laboratory conditions by exposition on treated potted plants. In addition to investigation of acute toxicity and persistence of harmful activity in both larvae and adults of A. bipunctata, demographic parameters were evaluated, to provide a comprehensive picture of the nontarget effects of these products. Field doses of the four insecticides caused detrimental effects to A. bipunctata; but in different ways. Overall, spinosad showed the best toxicological profile among the products tested. Emamectin benzoate could be considered a low-risk insecticide, but had high persistence. Chlorantraniliprole exhibited lethal effects on early instar larvae and adults, along with a long-lasting activity, instead spirotetramat showed a low impact on larval and adult mortality and can be considered a short-lived insecticide. However, demographic analysis demonstrated that chlorantraniliprole and spirotetramat caused sub-lethal effects. Our findings highlight that sole assessment of mortality can lead to underestimation of the full impact of pesticides on nontarget insects. Demographic analysis was demonstrated to be a sensitive method for detection of the sub-lethal effects of insecticides on A. bipunctata, and this approach should be considered for evaluation of insecticide selectivity. © The Author(s) 2017. Published by Oxford University Press on behalf of Entomological Society of America. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Experiences in therapy for lethal midline granuloma

International Nuclear Information System (INIS)

Tosaka, Kaoru; Ishikawa, Takeru

1982-01-01

Four cases of the lethal midline granuloma or malignant granuloma of the nose were treated by irradiation and chemotherapy, which are generally prescribed for malignant lymphomas. Clinical, histological and laboratory examination indicated that they were the lethal midline granuloma and clearly differentiated from Wegener's granulomatosis or malignant lymphoma. All of the cases exhibited primary remission. The four cases were observed up to 38, 22, 14, and 10 months since the beginning of the therapy, showing no local or general recurrence. (author)

Dysport: pharmacological properties and factors that influence toxin action.

Science.gov (United States)

Pickett, Andy

2009-10-01

The pharmacological properties of Dysport that influence toxin action are reviewed and compared with other botulinum toxin products. In particular, the subject of diffusion is examined and discussed based upon the evidence that currently exists, both from laboratory studies and from clinical data. Diffusion of botulinum toxin products is not related to the size of the toxin complex in the product since the complex dissociates under physiological conditions, releasing the naked neurotoxin to act. The active neurotoxin in Type A products is the same and therefore diffusion is equal when equal doses are administered.

Emergence of Escherichia coli encoding Shiga toxin 2f in human Shiga toxin-producing E-coli (STEC) infections in the Netherlands, January 2008 to December 2011

NARCIS (Netherlands)

Friesema, I.; van der Zwaluw, K.; Schuurman, T.; Kooistra-Smid, M.; Franz, E.; van Duynhoven, Y.; van Pelt, W.

2014-01-01

The Shiga toxins of Shiga toxin-producing Escherichia coli (STEC) can be divided into Shiga toxin 1 (Stx1) and Shiga toxin 2 (Stx2) with several sub-variants. Variant Stx(2f) is one of the latest described, but has been rarely associated with symptomatic human infections. In the enhanced STEC

Comparison of anorectic potencies of the trichothecenes T-2 toxin, HT-2 toxin and satratoxin G to the ipecac alkaloid emetine

Directory of Open Access Journals (Sweden)

Wenda Wu

2015-01-01

Full Text Available Trichothecene mycotoxins, potent translational inhibitors that are associated with human food poisonings and damp-building illnesses, are of considerable concern to animal and human health. Food refusal is a hallmark of exposure of experimental animals to deoxynivalenol (DON and other Type B trichothecenes but less is known about the anorectic effects of foodborne Type A trichothecenes (e.g., T-2 toxin, HT-2 toxin, airborne Type D trichothecenes (e.g., satratoxin G [SG] or functionally analogous metabolites that impair protein synthesis. Here, we utilized a well-described mouse model of food intake to compare the anorectic potencies of T-2 toxin, HT-2 toxin, and SG to that of emetine, a medicinal alkaloid derived from ipecac that inhibits translation. Intraperitoneal (IP administration with T-2 toxin, HT-2 toxin, emetine and SG evoked anorectic responses that occurred within 0.5 h that lasted up to 96, 96, 3 and 96 h, respectively, with lowest observed adverse effect levels (LOAELs being 0.1, 0.1, 2.5 and 0.25 mg/kg BW, respectively. When delivered via natural routes of exposure, T-2 toxin, HT-2 toxin, emetine (oral and SG (intranasal induced anorectic responses that lasted up to 48, 48, 3 and 6 h, respectively with LOAELs being 0.1, 0.1, 0.25, and 0.5 mg/kg BW, respectively. All four compounds were generally much more potent than DON which was previously observed to have LOAELs of 1 and 2.5 mg/kg BW after IP and oral dosing, respectively. Taken together, these anorectic potency data will be valuable in discerning the relative risks from trichothecenes and other translational inhibitors of natural origin.

How Parkinsonian Toxins Dysregulate the Autophagy Machinery

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Ruben K. Dagda

2013-11-01

Full Text Available Since their discovery, Parkinsonian toxins (6-hydroxydopamine, MPP+, paraquat, and rotenone have been widely employed as in vivo and in vitro chemical models of Parkinson’s disease (PD. Alterations in mitochondrial homeostasis, protein quality control pathways, and more recently, autophagy/mitophagy have been implicated in neurotoxin models of PD. Here, we highlight the molecular mechanisms by which different PD toxins dysregulate autophagy/mitophagy and how alterations of these pathways play beneficial or detrimental roles in dopamine neurons. The convergent and divergent effects of PD toxins on mitochondrial function and autophagy/mitophagy are also discussed in this review. Furthermore, we propose new diagnostic tools and discuss how pharmacological modulators of autophagy/mitophagy can be developed as disease-modifying treatments for PD. Finally, we discuss the critical need to identify endogenous and synthetic forms of PD toxins and develop efficient health preventive programs to mitigate the risk of developing PD.

On becoming neutral: effects of experimental neutralizing reconsidered.

Science.gov (United States)

van den Hout, M; van Pol, M; Peters, M

2001-12-01

Behaviour Research and Therapy 34 (1996) 889-898 found that writing out a negative thought produced anxiety and an urge to neutralize the thought, that instructing participants to neutralize the thought reduced anxiety/neutralization urge in the short run (i.e. within 2 min), but that in the control group 20 min without instruction was attended by the same reduction in anxiety/urge to neutralize ("natural decay"). The observations were made with pariticipants who scored high on "thought action fusion" and the experiment was set up as exerimental model of obsessions. We repeated the study with participants that were not selected on thought action fusion. All the findings reported by Behaviour Research and Therapy 34 (1996) 889-898 were replicated. Correlational analysis indicated that the strength of the effect was not related to scores on scales measuring "thought action fusion". Behaviour Research and Therapy 34 (1996) 889-898 did not assess whether non-neutralizing was followed by immediate reductions in distress. We did assess this and found that the larger part of the immediate reduction of distress after neutralization also occurs when no neutralization instruction is given. The effects of neutralization instructions in the present type of experiment are considerably less powerful than suggested earlier.

Dinophysis Toxins: Causative Organisms, Distribution and Fate in Shellfish

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Beatriz Reguera

2014-01-01

Full Text Available Several Dinophysis species produce diarrhoetic toxins (okadaic acid and dinophysistoxins and pectenotoxins, and cause gastointestinal illness, Diarrhetic Shellfish Poisoning (DSP, even at low cell densities (<103 cells·L−1. They are the main threat, in terms of days of harvesting bans, to aquaculture in Northern Japan, Chile, and Europe. Toxicity and toxin profiles are very variable, more between strains than species. The distribution of DSP events mirrors that of shellfish production areas that have implemented toxin regulations, otherwise misinterpreted as bacterial or viral contamination. Field observations and laboratory experiments have shown that most of the toxins produced by Dinophysis are released into the medium, raising questions about the ecological role of extracelular toxins and their potential uptake by shellfish. Shellfish contamination results from a complex balance between food selection, adsorption, species-specific enzymatic transformations, and allometric processes. Highest risk areas are those combining Dinophysis strains with high cell content of okadaates, aquaculture with predominance of mytilids (good accumulators of toxins, and consumers who frequently include mussels in their diet. Regions including pectenotoxins in their regulated phycotoxins will suffer from much longer harvesting bans and from disloyal competition with production areas where these toxins have been deregulated.

Immunization with Recombinant TcdB-Encapsulated Nanocomplex Induces Protection against Clostridium difficile Challenge in a Mouse Model

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Yi-Wen Liu

2017-07-01

Full Text Available Clostridium difficile is considered to be one of the major cause of infectious diarrhea in healthcare systems worldwide. Symptoms of C. difficile infection are caused largely by the production of two cytotoxins: toxin A (TcdA and toxin B (TcdB. Vaccine development is considered desirable as it would decrease the mounting medical costs and mortality associated with C. difficile infections. Biodegradable nanoparticles composed of poly-γ-glutamic acid (γ-PGA and chitosan have proven to be a safe and effective antigen delivery system for many viral vaccines. However, few studies have used this efficient antigen carrier for bacterial vaccine development. In this study, we eliminated the toxin activity domain of toxin B by constructing a recombinant protein rTcdB consists of residues 1852-2363 of TcdB receptor binding domain. The rTcdB was encapsulated in nanoparticles composed of γ-PGA and chitosan. Three rounds of intraperitoneal vaccination led to high anti-TcdB antibody responses and afforded mice full protection mice from lethal dose of C. difficile spore challenge. Protection was associated with high levels of toxin-neutralizing antibodies, and the rTcdB-encapsulated NPs elicited a longer-lasting antibody titers than antigen with the conventional adjuvant, aluminum hydroxide. Significant reductions in the level of proinflammatory cytokines and chemokines were observed in vaccinated mouse. These results suggested that polymeric nanocomplex-based vaccine design can be useful in developing vaccine against C. difficile infections.

ACTION OF DIPHTHERIA TOXIN IN THE GUINEA PIG

Science.gov (United States)

Baseman, Joel B.; Pappenheimer, A. M.; Gill, D. M.; Harper, Annabel A.

1970-01-01

The blood clearance and distribution in the tissues of 125I after intravenous injection of small doses (1.5–5 MLD or 0.08–0.25 µg) of 125I-labeled diphtheria toxin has been followed in guinea pigs and rabbits and compared with the fate of equivalent amounts of injected 125I-labeled toxoid and bovine serum albumin. Toxoid disappeared most rapidly from the blood stream and label accumulated and was retained in liver, spleen, and especially in kidney. Both toxin and BSA behaved differently. Label was found widely distributed among all the organs except the nervous system and its rate of disappearance from the tissues paralleled its disappearance from the circulation. There was no evidence for any particular affinity of toxin for muscle tissue or for a "target" organ. Previous reports by others that toxin causes specific and selective impairment of protein synthesis in muscle tissue were not confirmed. On the contrary, both in guinea pigs and rabbits, a reduced rate of protein synthesis was observed in all tissues that had taken up the toxin label. In tissues removed from intoxicated animals of both species there was an associated reduction in aminoacyl transferase 2 content. It is concluded that the primary action of diphtheria toxin in the living animal is to effect the inactivation of aminoacyl transferase 2. The resulting inhibition in rate of protein synthesis leads to morphologic damage in all tissues reached by the toxin and ultimately to death of the animal. PMID:5511567

Back to the future: revisiting HIV-1 lethal mutagenesis

Science.gov (United States)

Dapp, Michael J.; Patterson, Steven E.; Mansky, Louis M.

2012-01-01

The concept of eliminating HIV-1 infectivity by elevating the viral mutation rate was first proposed over a decade ago, even though the general concept had been conceived earlier for RNA viruses. Lethal mutagenesis was originally viewed as a novel chemotherapeutic approach for treating HIV-1 infection in which use of a viral mutagen would over multiple rounds of replication lead to the lethal accumulation of mutations, rendering the virus population non infectious – known as the slow mutation accumulation model. There have been limitations in obtaining good efficacy data with drug leads, leaving some doubt into clinical translation. More recent studies of the APOBEC3 proteins as well as new progress in the use of nucleoside analogs for inducing lethal mutagenesis have helped to refocus attention on rapid induction of HIV-1 lethal mutagenesis in a single or limited number of replication cycles leading to a rapid mutation accumulation model. PMID:23195922

Comparison of T-2 Toxin and HT-2 Toxin Distributed in the Skeletal System with That in Other Tissues of Rats by Acute Toxicity Test.

Science.gov (United States)

Yu, Fang Fang; Lin, Xia Lu; Yang, Lei; Liu, Huan; Wang, Xi; Fang, Hua; Lammi, ZMikko J; Guo, Xiong

2017-11-01

Twelve healthy rats were divided into the T-2 toxin group receiving gavage of 1 mg/kg T-2 toxin and the control group receiving gavage of normal saline. Total relative concentrations of T-2 toxin and HT-2 toxin in the skeletal system (thighbone, knee joints, and costal cartilage) were significantly higher than those in the heart, liver, and kidneys (P skeletal system (thighbone and costal cartilage) were also significantly higher than those in the heart, liver, and kidneys. The rats administered T-2 toxin showed rapid metabolism compared with that in rats administered HT-2 toxin, and the metabolic conversion rates in the different tissues were 68.20%-90.70%. Copyright © 2017 The Editorial Board of Biomedical and Environmental Sciences. Published by China CDC. All rights reserved.

Perinatal-lethal Gaucher disease presenting as hydrops fetalis.

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BenHamida, Emira; Ayadi, Imene; Ouertani, Ines; Chammem, Maroua; Bezzine, Ahlem; BenTmime, Riadh; Attia, Leila; Mrad, Ridha; Marrakchi, Zahra

2015-01-01

Perinatal-lethal Gaucher disease is very rare and is considered a variant of type 2 Gaucher disease that occurs in the neonatal period. The most distinct features of perinatal-lethal Gaucher disease are non-immune hydrops fetalis. Less common signs of the disease are hepatosplenomegaly, ichthyosis and arthrogryposis. We report a case of Gaucher's disease (type 2) diagnosed in a newborn who presented with Hydrops Fetalis.

Crystal structure of Clostridium difficile toxin A

Energy Technology Data Exchange (ETDEWEB)

Chumbler, Nicole M.; Rutherford, Stacey A.; Zhang, Zhifen; Farrow, Melissa A.; Lisher, John P.; Farquhar, Erik; Giedroc, David P.; Spiller, Benjamin W.; Melnyk, Roman A.; Lacy, D. Borden

2016-01-11

Clostridium difficile infection is the leading cause of hospital-acquired diarrhoea and pseudomembranous colitis. Disease is mediated by the actions of two toxins, TcdA and TcdB, which cause the diarrhoea, as well as inflammation and necrosis within the colon. The toxins are large (308 and 270 kDa, respectively), homologous (47% amino acid identity) glucosyltransferases that target small GTPases within the host. The multidomain toxins enter cells by receptor-mediated endocytosis and, upon exposure to the low pH of the endosome, insert into and deliver two enzymatic domains across the membrane. Eukaryotic inositol-hexakisphosphate (InsP6) binds an autoprocessing domain to activate a proteolysis event that releases the N-terminal glucosyltransferase domain into the cytosol. Here, we report the crystal structure of a 1,832-amino-acid fragment of TcdA (TcdA₁₈₃₂), which reveals a requirement for zinc in the mechanism of toxin autoprocessing and an extended delivery domain that serves as a scaffold for the hydrophobic α-helices involved in pH-dependent pore formation. A surface loop of the delivery domain whose sequence is strictly conserved among all large clostridial toxins is shown to be functionally important, and is highlighted for future efforts in the development of vaccines and novel therapeutics.

Botulinum Toxin and Muscle Atrophy: A Wanted or Unwanted Effect.

Science.gov (United States)

Durand, Paul D; Couto, Rafael A; Isakov, Raymond; Yoo, Donald B; Azizzadeh, Babak; Guyuron, Bahman; Zins, James E

2016-04-01

While the facial rejuvenating effect of botulinum toxin type A is well known and widespread, its use in body and facial contouring is less common. We first describe its use for deliberate muscle volume reduction, and then document instances of unanticipated and undesirable muscle atrophy. Finally, we investigate the potential long-term adverse effects of botulinum toxin-induced muscle atrophy. Although the use of botulinum toxin type A in the cosmetic patient has been extensively studied, there are several questions yet to be addressed. Does prolonged botulinum toxin treatment increase its duration of action? What is the mechanism of muscle atrophy and what is the cause of its reversibility once treatment has stopped? We proceed to examine how prolonged chemodenervation with botulinum toxin can increase its duration of effect and potentially contribute to muscle atrophy. Instances of inadvertent botulinum toxin-induced atrophy are also described. These include the "hourglass deformity" secondary to botulinum toxin type A treatment for migraine headaches, and a patient with atrophy of multiple facial muscles from injections for hemifacial spasm. Numerous reports demonstrate that muscle atrophy after botulinum toxin type A treatment occurs and is both reversible and temporary, with current literature supporting the notion that repeated chemodenervation with botulinum toxin likely responsible for both therapeutic and incidental temporary muscle atrophy. Furthermore, duration of response may be increased with subsequent treatments, thus minimizing frequency of reinjection. Practitioners should be aware of the temporary and reversible effect of botulinum toxin-induced muscle atrophy and be prepared to reassure patients on this matter. © 2016 The American Society for Aesthetic Plastic Surgery, Inc. Reprints and permission: journals.permissions@oup.com.

Oxidative Stress in Shiga Toxin Production by Enterohemorrhagic Escherichia coli

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Katarzyna Licznerska

2016-01-01

Full Text Available Virulence of enterohemorrhagic Escherichia coli (EHEC strains depends on production of Shiga toxins. These toxins are encoded in genomes of lambdoid bacteriophages (Shiga toxin-converting phages, present in EHEC cells as prophages. The genes coding for Shiga toxins are silent in lysogenic bacteria, and prophage induction is necessary for their efficient expression and toxin production. Under laboratory conditions, treatment with UV light or antibiotics interfering with DNA replication are commonly used to induce lambdoid prophages. Since such conditions are unlikely to occur in human intestine, various research groups searched for other factors or agents that might induce Shiga toxin-converting prophages. Among other conditions, it was reported that treatment with H2O2 caused induction of these prophages, though with efficiency significantly lower relative to UV-irradiation or mitomycin C treatment. A molecular mechanism of this phenomenon has been proposed. It appears that the oxidative stress represents natural conditions provoking induction of Shiga toxin-converting prophages as a consequence of H2O2 excretion by either neutrophils in infected humans or protist predators outside human body. Finally, the recently proposed biological role of Shiga toxin production is described in this paper, and the “bacterial altruism” and “Trojan Horse” hypotheses, which are connected to the oxidative stress, are discussed.

Botulinum Toxin in Neurogenic Detrusor Overactivity

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Carlos Arturo Levi D'Ancona

2012-09-01

Full Text Available Purpose To evaluate the effects of botulinum toxin on urodynamic parameters and quality of life in patients with neurogenic detrusor overactivity. Methods Thirty four adult patients with spinal cord injury and detrusor overactivity were selected. The patients received 300 units of botulinum toxin type A. The endpoints evaluated with the episodes of urinary incontinence and measured the maximum cystometric capacity, maximum amplitude of detrusor pressure and bladder compliance at the beginning and end of the study (24 weeks and evaluated the quality of life by applying the Qualiveen questionnaire. Results A significant decrease in the episodes of urinary incontinence was observed. All urodynamic parameters presented a significant improvement. The same was observed in the quality of life index and the specific impact of urinary problems scores from the Qualiveen questionnaire. Six patients did not complete the study, two due to incomplete follow-up, and four violated protocol and were excluded from the analyses. No systemic adverse events of botulinum toxin type A were reported. Conclusions A botulinum toxin type A showed a significantly improved response in urodynamics parameters and specific and general quality of life.

Bacterial toxin-antitoxin systems: more than selfish entities?

OpenAIRE

Laurence Van Melderen; Manuel Saavedra De Bast

2009-01-01

Bacterial toxin?antitoxin (TA) systems are diverse and widespread in the prokaryotic kingdom. They are composed of closely linked genes encoding a stable toxin that can harm the host cell and its cognate labile antitoxin, which protects the host from the toxin's deleterious effect. TA systems are thought to invade bacterial genomes through horizontal gene transfer. Some TA systems might behave as selfish elements and favour their own maintenance at the expense of their host. As a consequence,...

125I-induced DNA double strand breaks: use in calibration of the neutral filter elution technique and comparison with X-ray induced breaks

International Nuclear Information System (INIS)

Radford, I.R.; Hodgson, G.S.

1985-01-01

The neutral filter elution assay, for measurement of DNA double strand breakage, has been calibrated using mouse L cells and Chinese hamster V79 cells labelled with [ 125 I]dUrd and then held at liquid nitrogen temperature to accumulate decays. The basis of the calibration is the observation that each 125 I decay, occurring in DNA, produces a DNA double strand break. Linear relationships between 125 I decays per cell and lethal lesions per cell (minus natural logarithm survival) and the level of elution, were found. Using the calibration data, it was calculated that the yield of DNA double strand breaks after X-irradiation of both cell types was from 6 to 9 x 10 -12 DNA double strand breaks per Gy per dalton of DNA, for doses greater than 6 Gy. Neutral filter elution and survival data for X-irradiated and 125 I-labelled cells suggested that the relationships between lethal lesions and DNA double strand breakage were significantly different for both cell types. An attempt was made to study the repair kinetics for 125 I-induced DNA double strand breaks, but was frustrated by the rapid DNA degradation which occurs in cells that have been killed by the freezing-thawing process. (author)

Gene therapy for carcinoma of the breast: Genetic toxins

International Nuclear Information System (INIS)

Vassaux, Georges; Lemoine, Nick R

2000-01-01

Gene therapy was initially envisaged as a potential treatment for genetically inherited, monogenic disorders. The applications of gene therapy have now become wider, however, and include cardiovascular diseases, vaccination and cancers in which conventional therapies have failed. With regard to oncology, various gene therapy approaches have been developed. Among them, the use of genetic toxins to kill cancer cells selectively is emerging. Two different types of genetic toxins have been developed so far: the metabolic toxins and the dominant-negative class of toxins. This review describes these two different approaches, and discusses their potential applications in cancer gene therapy

Modification of opiate agonist binding by pertussis toxin

Energy Technology Data Exchange (ETDEWEB)

Abood, M.E.; Lee, N.M.; Loh, H.H.

1986-03-05

Opiate agonist binding is decreased by GTP, suggesting the possible involvement of GTP binding proteins in regulation of opiate receptor binding. This possibility was addressed by asking whether pertussis toxin treatment, which results in ADP-ribosylation and modification of G proteins, would alter opiate agonist binding. The striatum was chosen for the initial brain area to be studied, since regulation of opiate action in this area had been shown to be modified by pertussis toxin. Treatment of striatal membranes with pertussis toxin results in up to a 55% decrease in /sup 3/(H)-DADLE binding as compared with membranes treated identically without toxin. This corresponds to a near complete ADP-ribosylation of both G proteins in the striatal membrane. The decrease in agonist binding appears to be due to an altered affinity of the receptor for agonist as opposed to a decrease in the number of sites. This effect of pertussis toxin on opiate agonist binding demonstrates the actual involvement of G proteins in regulation of opiate receptor binding.

Modification of opiate agonist binding by pertussis toxin

International Nuclear Information System (INIS)

Abood, M.E.; Lee, N.M.; Loh, H.H.

1986-01-01

Opiate agonist binding is decreased by GTP, suggesting the possible involvement of GTP binding proteins in regulation of opiate receptor binding. This possibility was addressed by asking whether pertussis toxin treatment, which results in ADP-ribosylation and modification of G proteins, would alter opiate agonist binding. The striatum was chosen for the initial brain area to be studied, since regulation of opiate action in this area had been shown to be modified by pertussis toxin. Treatment of striatal membranes with pertussis toxin results in up to a 55% decrease in 3 (H)-DADLE binding as compared with membranes treated identically without toxin. This corresponds to a near complete ADP-ribosylation of both G proteins in the striatal membrane. The decrease in agonist binding appears to be due to an altered affinity of the receptor for agonist as opposed to a decrease in the number of sites. This effect of pertussis toxin on opiate agonist binding demonstrates the actual involvement of G proteins in regulation of opiate receptor binding

ADP-ribosylation of transducin by pertussis toxin

International Nuclear Information System (INIS)

Watkins, P.A.; Burns, D.L.; Kanaho, Y.; Liu, T.Y.; Hewlett, E.L.; Moss, J.

1985-01-01

Transducin, the guanyl nucleotide-binding regulatory protein of retinal rod outer segments that couples the photon receptor, rhodopsin, with the light-activated cGMP phosphodiesterase, can be resolved into two functional components, T alpha and T beta gamma. T alpha (39 kDa), which is [ 32 P]ADP-ribosylated by pertussis toxin and [ 32 P]NAD in rod outer segments and in purified transducin, was also labeled by the toxin after separation from T beta gamma (36 kDa and approximately 10 kDa); neither component of T beta gamma was a pertussis toxin substrate. Labeling of T alpha was enhanced by T beta gamma and was maximal at approximately 1:1 molar ratio of T alpha : T beta gamma. Limited proteolysis by trypsin of T alpha in the presence of guanyl-5'-yl imidodiphosphate (Gpp(NH)p) resulted in the sequential appearance of proteins of 38 and 32 kDa. The amino terminus of both 38- and 32 -kDa proteins was leucine, whereas that of T alpha could not be identified and was assumed to be blocked. The 32 -kDa peptide was not a pertussis toxin substrate. Labeling of the 38-kDa protein was poor and was not enhanced by T beta gamma. Trypsin treatment of [ 32 P]ADP-ribosyl-T alpha produced a labeled 37-38-kDa doublet followed by appearance of radioactivity at the dye front. It appears, therefore, that, although the 38-kDa protein was poor toxin substrate, it contained the ADP-ribosylation site. Without rhodopsin, labeling of T alpha (in the presence of T beta gamma) was unaffected by Gpp(NH)p, guanosine 5'-O-(thiotriphosphate) (GTP gamma S), GTP, GDP, and guanosine 5'-O-(thiodiphosphate) (GDP beta S) but was increased by ATP. When photolyzed rhodopsin and T beta gamma were present, Gpp(NH)p and GTP gamma S decreased [ 32 P]ADP-ribosylation by pertussis toxin. Thus, pertussis toxin-catalyzed [ 32 P]ADP-ribosylation of T alpha was affected by nucleotides, rhodopsin and light in addition to T beta gamma

Marine toxins and their toxicological significance: An overview

Digital Repository Service at National Institute of Oceanography (India)

Sarkar, A.

, Hemolysins-1 and hemolysin-2, saxitoxin, neosaxitoxin, gonyautoxin, tetrodotoxin, ptychodiscus brevis toxin and theonellamide F. According to their mode of action, these toxins are classified into different categories such as cytotoxin, enterotoxin...

Toxin synergism in snake venoms

DEFF Research Database (Denmark)

Laustsen, Andreas Hougaard

2016-01-01

Synergism between venom toxins exists for a range of snake species. Synergism can be derived from both intermolecular interactions and supramolecular interactions between venom components, and can be the result of toxins targeting the same protein, biochemical pathway or physiological process. Few...... simple systematic tools and methods for determining the presence of synergism exist, but include co-administration of venom components and assessment of Accumulated Toxicity Scores. A better understanding of how to investigate synergism in snake venoms may help unravel strategies for developing novel...

Lethal neonatal short-limbed dwarfism

International Nuclear Information System (INIS)

Kim, Ok Hwa; Yim, Chung Ik; Bahk, Yong Whee

1986-01-01

We have detailed our experiences on 6 cases of neonatal lethal short-limbed dwarfism and reviewed the articles. They include, achondrogenesis, thanatophoric dysplasia, asphyxiating thoracic dysplasia, osteogenesis imperfect a congenita, and hypophosphatasia lethals. Five babies were born alive but died soon after birth and one was a stillbirth. The main cause of failure to thrive was respiratory insufficiency. Each case was having quite characteristic radiologic findings, even if the general appearances were similar to the achondroplasts clinically. Precise diagnosis is very important for genetic counselling of the parents and alarm to them the possibility of bone dysplasias to the next offsprings. For this purpose, the radiologists play major role for the correct diagnosis. We stress that when the baby is born with short-limbed dwarfism, whole body radiogram should be taken including lateral view and postmortem radiogram is also very precious.

Lethal neonatal short-limbed dwarfism

Energy Technology Data Exchange (ETDEWEB)

Kim, Ok Hwa; Yim, Chung Ik; Bahk, Yong Whee [Catholic Medical College, Seoul (Korea, Republic of)

1986-02-15

We have detailed our experiences on 6 cases of neonatal lethal short-limbed dwarfism and reviewed the articles. They include, achondrogenesis, thanatophoric dysplasia, asphyxiating thoracic dysplasia, osteogenesis imperfect a congenita, and hypophosphatasia lethals. Five babies were born alive but died soon after birth and one was a stillbirth. The main cause of failure to thrive was respiratory insufficiency. Each case was having quite characteristic radiologic findings, even if the general appearances were similar to the achondroplasts clinically. Precise diagnosis is very important for genetic counselling of the parents and alarm to them the possibility of bone dysplasias to the next offsprings. For this purpose, the radiologists play major role for the correct diagnosis. We stress that when the baby is born with short-limbed dwarfism, whole body radiogram should be taken including lateral view and postmortem radiogram is also very precious.

Toxin-independent virulence of Bacillus anthracis in rabbits.

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Haim Levy

Full Text Available The accepted paradigm states that anthrax is both an invasive and toxinogenic disease and that the toxins play a major role in pathogenicity. In the guinea pig (GP model we have previously shown that deletion of all three toxin components results in a relatively moderate attenuation in virulence, indicating that B. anthracis possesses an additional toxin-independent virulence mechanism. To characterize this toxin-independent mechanism in anthrax disease, we developed a new rabbit model by intravenous injection (IV of B. anthracis encapsulated vegetative cells, artificially creating bacteremia. Using this model we were able to demonstrate that also in rabbits, B. anthracis mutants lacking the toxins are capable of killing the host within 24 hours. This virulent trait depends on the activity of AtxA in the presence of pXO2, as, in the absence of the toxin genes, deletion of either component abolishes virulence. Furthermore, this IV virulence depends mainly on AtxA rather than the whole pXO1. A similar pattern was shown in the GP model using subcutaneous (SC administration of spores of the mutant strains, demonstrating the generality of the phenomenon. The virulent strains showed higher bacteremia levels and more efficient tissue dissemination; however our interpretation is that tissue dissemination per se is not the main determinant of virulence whose exact nature requires further elucidation.

Radiation resistance of paralytic shellfish poison (PSP) toxins

Energy Technology Data Exchange (ETDEWEB)

San Juan, Edith M

2000-04-01

Radiation resistance of paralytic shellfish poison (PSP) toxins, obtained from Pyrodinium bahamense var. compressum in shellstocks of green mussels, was determined by subjecting the semi-purified toxin extract as well as the shellstocks of green mussels to high doses of ionizing radiation of 5, 10, 15 and 20 kGy. The concentration of the PSP toxins was determined by the Standard Mouse Bioassay (SMB) method. The radiation assistance of the toxins was determined by plotting the PSP toxin concentration versus applied dose in a semilog paper. The D{sub 10} value or decimal reduction dose was obtained from the straight line which is the dose required to reduce the toxicity level by 90%. The effects of irradiation on the quality of green mussels in terms of its physico-chemical, microbiological and sensory attributes were also conducted. The effect of irradiation on the fatty acid components of green mussels was determined by gas chromatography. Radiation resistance of the PSP toxins was determined to be lower in samples with initially high toxicity level as compared with samples with initially low toxicity level. The D{sub 10} values of samples with initially high PSP level were 28.5 kGy in shellstocks of green musssels and 17.5 kGy in the semi-purified toxin extract. When the PSP level was low initially, the D{sub 10} values were as high as 57.5 and 43.5 kGy in shellstocks of green mussels for the two trials, and 43.0 kGy in semi-purified toxin extract. The microbial load of the irradiated mussels was remarkably reduced. No differnce in color and odor characteristics were observed in the mussel samples subjected to varying doses of ionizing radiation. There was darkening in the color of mussel meat and its juice. The concentration of the fatty acid components in the fresh green mussels were considerably higher as compared with those present in the irradiated mussels, though some volatile fatty acids were detected as a result of irradiation. (Author)

Radiation resistance of paralytic shellfish poison (PSP) toxins

International Nuclear Information System (INIS)

San Juan, Edith M.

2000-04-01

Radiation resistance of paralytic shellfish poison (PSP) toxins, obtained from Pyrodinium bahamense var. compressum in shellstocks of green mussels, was determined by subjecting the semi-purified toxin extract as well as the shellstocks of green mussels to high doses of ionizing radiation of 5, 10, 15 and 20 kGy. The concentration of the PSP toxins was determined by the Standard Mouse Bioassay (SMB) method. The radiation assistance of the toxins was determined by plotting the PSP toxin concentration versus applied dose in a semilog paper. The D 10 value or decimal reduction dose was obtained from the straight line which is the dose required to reduce the toxicity level by 90%. The effects of irradiation on the quality of green mussels in terms of its physico-chemical, microbiological and sensory attributes were also conducted. The effect of irradiation on the fatty acid components of green mussels was determined by gas chromatography. Radiation resistance of the PSP toxins was determined to be lower in samples with initially high toxicity level as compared with samples with initially low toxicity level. The D 10 values of samples with initially high PSP level were 28.5 kGy in shellstocks of green musssels and 17.5 kGy in the semi-purified toxin extract. When the PSP level was low initially, the D 10 values were as high as 57.5 and 43.5 kGy in shellstocks of green mussels for the two trials, and 43.0 kGy in semi-purified toxin extract. The microbial load of the irradiated mussels was remarkably reduced. No differnce in color and odor characteristics were observed in the mussel samples subjected to varying doses of ionizing radiation. There was darkening in the color of mussel meat and its juice. The concentration of the fatty acid components in the fresh green mussels were considerably higher as compared with those present in the irradiated mussels, though some volatile fatty acids were detected as a result of irradiation. (Author)

Bacterial community affects toxin production by Gymnodinium catenatum.

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Maria E Albinsson

Full Text Available The paralytic shellfish toxin (PST-producing dinoflagellate Gymnodinium catenatum grows in association with a complex marine bacterial community that is both essential for growth and can alter culture growth dynamics. Using a bacterial community replacement approach, we examined the intracellular PST content, production rate, and profile of G. catenatum cultures grown with bacterial communities of differing complexity and composition. Clonal offspring were established from surface-sterilized resting cysts (produced by sexual crosses of strain GCDE06 and strain GCLV01 and grown with: 1 complex bacterial communities derived from each of the two parent cultures; 2 simplified bacterial communities composed of the G. catenatum-associated bacteria Marinobacter sp. strain DG879 or Alcanivorax sp. strain DG881; 3 a complex bacterial community associated with an untreated, unsterilized sexual cross of the parents. Toxin content (STX-equivalent per cell of clonal offspring (134-197 fmol STX cell(-1 was similar to the parent cultures (169-206 fmol STX cell(-1, however cultures grown with single bacterial types contained less toxin (134-146 fmol STX cell(-1 than offspring or parent cultures grown with more complex mixed bacterial communities (152-176 fmol STX cell(-1. Specific toxin production rate (fmol STX day(-1 was strongly correlated with culture growth rate. Net toxin production rate (fmol STX cell(-1 day(-1 did not differ among treatments, however, mean net toxin production rate of offspring was 8-fold lower than the parent cultures, suggesting that completion of the sexual lifecycle in laboratory cultures leads to reduced toxin production. The PST profiles of offspring cultures were most similar to parent GCDE06 with the exception of cultures grown with Marinobacter sp. DG879 which produced higher proportions of dcGTX2+3 and GC1+2, and lower proportions of C1+2 and C3+4. Our data demonstrate that the bacterial community can alter intracellular STX

Bacterial community affects toxin production by Gymnodinium catenatum.

Science.gov (United States)

Albinsson, Maria E; Negri, Andrew P; Blackburn, Susan I; Bolch, Christopher J S

2014-01-01

The paralytic shellfish toxin (PST)-producing dinoflagellate Gymnodinium catenatum grows in association with a complex marine bacterial community that is both essential for growth and can alter culture growth dynamics. Using a bacterial community replacement approach, we examined the intracellular PST content, production rate, and profile of G. catenatum cultures grown with bacterial communities of differing complexity and composition. Clonal offspring were established from surface-sterilized resting cysts (produced by sexual crosses of strain GCDE06 and strain GCLV01) and grown with: 1) complex bacterial communities derived from each of the two parent cultures; 2) simplified bacterial communities composed of the G. catenatum-associated bacteria Marinobacter sp. strain DG879 or Alcanivorax sp. strain DG881; 3) a complex bacterial community associated with an untreated, unsterilized sexual cross of the parents. Toxin content (STX-equivalent per cell) of clonal offspring (134-197 fmol STX cell(-1)) was similar to the parent cultures (169-206 fmol STX cell(-1)), however cultures grown with single bacterial types contained less toxin (134-146 fmol STX cell(-1)) than offspring or parent cultures grown with more complex mixed bacterial communities (152-176 fmol STX cell(-1)). Specific toxin production rate (fmol STX day(-1)) was strongly correlated with culture growth rate. Net toxin production rate (fmol STX cell(-1) day(-1)) did not differ among treatments, however, mean net toxin production rate of offspring was 8-fold lower than the parent cultures, suggesting that completion of the sexual lifecycle in laboratory cultures leads to reduced toxin production. The PST profiles of offspring cultures were most similar to parent GCDE06 with the exception of cultures grown with Marinobacter sp. DG879 which produced higher proportions of dcGTX2+3 and GC1+2, and lower proportions of C1+2 and C3+4. Our data demonstrate that the bacterial community can alter intracellular STX

Diphtheria toxin-induced channels in Vero cells selective for monovalent cations

International Nuclear Information System (INIS)

Sandvig, K.; Olsnes, S.

1988-01-01

Ion fluxes associated with translocation of diphtheria toxin across the surface membrane of Vero cells were studied. When cells with surface-bound toxin were exposed to low pH to induce toxin entry, the cells became permeable to Na+, K+, H+, choline+, and glucosamine+. There was no increased permeability to Cl-, SO4(-2), glucose, or sucrose, whereas the uptake of 45 Ca2+ was slightly increased. The influx of Ca2+, which appears to be different from that of monovalent cations, was reduced by several inhibitors of anion transport and by verapamil, Mn2+, Co2+, and Ca2+, but not by Mg2+. The toxin-induced fluxes of N+, K+, and protons were inhibited by Cd2+. Cd2+ also protected the cells against intoxication by diphtheria toxin, suggesting that the open cation-selective channel is required for toxin translocation. The involvement of the toxin receptor is discussed

Cholix Toxin, a Novel ADP-ribosylating Factor from Vibrio cholerae

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Jorgensen, Rene; Purdy, Alexandra E.; Fieldhouse, Robert J.; Kimber, Matthew S.; Bartlett, Douglas H.; Merrill, A. Rod (Guelph); (NIH); (UCSD)

2008-07-15

The ADP-ribosyltransferases are a class of enzymes that display activity in a variety of bacterial pathogens responsible for causing diseases in plants and animals, including those affecting mankind, such as diphtheria, cholera, and whooping cough. We report the characterization of a novel toxin from Vibrio cholerae, which we call cholix toxin. The toxin is active against mammalian cells (IC50 = 4.6 {+-} 0.4 ng/ml) and crustaceans (Artemia nauplii LD50 = 10 {+-} 2 {mu}g/ml). Here we show that this toxin is the third member of the diphthamide-specific class of ADP-ribose transferases and that it possesses specific ADP-ribose transferase activity against ribosomal eukaryotic elongation factor 2. We also describe the high resolution crystal structures of the multidomain toxin and its catalytic domain at 2.1- and 1.25-{angstrom} resolution, respectively. The new structural data show that cholix toxin possesses the necessary molecular features required for infection of eukaryotes by receptor-mediated endocytosis, translocation to the host cytoplasm, and inhibition of protein synthesis by specific modification of elongation factor 2. The crystal structures also provide important insight into the structural basis for activation of toxin ADP-ribosyltransferase activity. These results indicate that cholix toxin may be an important virulence factor of Vibrio cholerae that likely plays a significant role in the survival of the organism in an aquatic environment.

Rabies Virus Antibodies from Oral Vaccination as a Correlate of Protection against Lethal Infection in Wildlife

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Susan M. Moore

2017-07-01

Full Text Available Both cell-mediated and humoral immune effectors are important in combating rabies infection, although the humoral response receives greater attention regarding rabies prevention. The principle of preventive vaccination has been adopted for strategies of oral rabies vaccination (ORV of wildlife reservoir populations for decades to control circulation of rabies virus in free-ranging hosts. There remains much debate about the levels of rabies antibodies (and the assays to measure them that confer resistance to rabies virus. In this paper, data from published literature and our own unpublished animal studies on the induction of rabies binding and neutralizing antibodies following oral immunization of animals with live attenuated or recombinant rabies vaccines, are examined as correlates of protection against lethal rabies infection in captive challenge settings. Analysis of our studies suggests that, though serum neutralization test results are expected to reflect in vivo protection, the blocking enzyme linked immunosorbent assay (ELISA result at Day 28 was a better predictor of survival. ELISA kits may have an advantage of greater precision and ability to compare results among different studies and laboratories based on the inherent standardization of the kit format. This paper examines current knowledge and study findings to guide meaningful interpretation of serology results in oral baiting monitoring.

Botulinum toxin type A versus botulinum toxin type B for cervical dystonia.

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Duarte, Gonçalo S; Castelão, Mafalda; Rodrigues, Filipe B; Marques, Raquel E; Ferreira, Joaquim; Sampaio, Cristina; Moore, Austen P; Costa, João

2016-10-26

This is an update of a Cochrane review first published in 2003. Cervical dystonia is the most common form of focal dystonia and is a disabling disorder characterised by painful involuntary head posturing. There are two available formulations of botulinum toxin, with botulinum toxin type A (BtA) usually considered the first line therapy for this condition. Botulinum toxin type B (BtB) is an alternative option, with no compelling theoretical reason why it might not be as- or even more effective - than BtA. To compare the efficacy, safety and tolerability of botulinum toxin type A (BtA) versus botulinum toxin type B (BtB) in people with cervical dystonia. To identify studies for this review we searched the Cochrane Movement Disorders Group Trials Register, the Cochrane Central Register of Controlled Trials (CENTRAL), MEDLINE, Embase, reference lists of articles and conference proceedings. All elements of the search, with no language restrictions, were last run in October 2016. Double-blind, parallel, randomised, placebo-controlled trials (RCTs) comparing BtA versus BtB in adults with cervical dystonia. Two independent authors assessed records, selected included studies, extracted data using a paper pro forma, and evaluated the risk of bias. We resolved disagreements by consensus or by consulting a third author. We performed meta-analyses using the random-effects model, for the comparison BtA versus BtB to estimate pooled effects and corresponding 95% confidence intervals (95% CI). No prespecified subgroup analyses were carried out. The primary efficacy outcome was improvement on any validated symptomatic rating scale, and the primary safety outcome was the proportion of participants with adverse events. We included three RCTs, all new to this update, of very low to low methodological quality, with a total of 270 participants.Two studies exclusively enrolled participants with a known positive response to BtA treatment. This raises concerns of population enrichment

[Botulism: structure and function of botulinum toxin and its clinical application].

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Oguma, Keiji; Yamamoto, Yumiko; Suzuki, Tomonori; Fatmawati, Ni Nengah Dwi; Fujita, Kumiko

2012-08-01

Clostridium botulinum produces seven immunological distinct poisonous neurotoxins, A to G, with molecular masses of approximately 150kDa. In acidic foods and culture fluid, the neurotoxins associate with non-toxic components, and form large complexes designated progenitor toxins. The progenitor toxins are found in three forms named LL, L, and M. These neurotoxins and progenitor toxins were purified, and whole nucleotide sequences of their structure genes were determined. In this manuscript, the structure and function of these toxins, and the application of these toxins to clinical usage have been described.

Botulinum toxin in the treatment of vocal fold nodules.

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Allen, Jacqui E; Belafsky, Peter C

2009-12-01

Promising new techniques in the management of vocal fold nodules have been developed in the past 2 years. Simultaneously, the therapeutic use of botulinum toxin has rapidly expanded. This review explores the use of botulinum toxin in treatment of vocal nodules and summarizes current therapeutic concepts. New microsurgical instruments and techniques, refinements in laser technology, radiosurgical excision and steroid intralesional injections are all promising new techniques in the management of vocal nodules. Botulinum toxin-induced 'voice rest' is a new technique we have employed in patients with recalcitrant nodules. Successful resolution of nodules is possible with this technique, without the risk of vocal fold scarring inherent in dissection/excision techniques. Botulinum toxin usage is exponentially increasing, and large-scale, long-term studies demonstrate its safety profile. Targeted vocal fold temporary paralysis induced by botulinum toxin injection is a new, well tolerated and efficacious treatment in patients with persistent vocal fold nodules.

Synthesis of protein in intestinal cells exposed to cholera toxin

International Nuclear Information System (INIS)

Peterson, J.W.; Berg, W.D. Jr.; Coppenhaver, D.H.

1987-01-01

The mechanism by which cyclic adenosine monophosphate (AMP), formed by intestinal epithelial cells in response to cholera toxin, ultimately results in alterations in water and electrolyte transport is poorly understood. Several studies have indicated that inhibitors of transcription or translation block much of the transport of ions and water in the intestine and edema formation in tissue elicited by cholera toxin. Data presented in this study confirmed the inhibitory effects of cycloheximide on cholera toxin-induced fluid accumulation in the rabbit intestinal loop model. Neither cycloheximide nor actinomycin D altered the amount of cyclic AMP that accumulated in intestinal cells and Chinese hamster ovary cells exposed to cholera toxin. An increase in [ 3 H] leucine incorporation was readily demonstrable in intestinal epithelial cells from rabbits challenged with Vibrio cholerae. Similarly, intestinal epithelial cells incubated with cholera toxin for 4 hr synthesized substantially more protein than controls as determined by relative incorporation of [ 35 S] methionine. Most of the new protein synthesized in response to cholera toxin was membrane associated and of high molecular weight. The possible significance of the toxin-induced protein relative to cholera pathogenesis was discussed

Botulinum Toxin in Management of Limb Tremor

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Elina Zakin

2017-11-01

Full Text Available Essential tremor is characterized by persistent, usually bilateral and symmetric, postural or kinetic activation of agonist and antagonist muscles involving either the distal or proximal upper extremity. Quality of life is often affected and one’s ability to perform daily tasks becomes impaired. Oral therapies, including propranolol and primidone, can be effective in the management of essential tremor, although adverse effects can limit their use and about 50% of individuals lack response to oral pharmacotherapy. Locally administered botulinum toxin injection has become increasingly useful in the management of essential tremor. Targeting of select muscles with botulinum toxin is an area of active research, and muscle selection has important implications for toxin dosing and functional outcomes. The use of anatomical landmarks with palpation, EMG guidance, electrical stimulation, and ultrasound has been studied as a technique for muscle localization in toxin injection. Earlier studies implemented a standard protocol for the injection of (predominantly wrist flexors and extensors using palpation and EMG guidance. Targeting of muscles by selection of specific activators of tremor (tailored to each patient using kinematic analysis might allow for improvement in efficacy, including functional outcomes. It is this individualized muscle selection and toxin dosing (requiring injection within various sites of a single muscle that has allowed for success in the management of tremors.

Hemagglutinin-specific neutralization of subacute sclerosing panencephalitis viruses.

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Miguel Ángel Muñoz-Alía

Full Text Available Subacute sclerosing panencephalitis (SSPE is a progressive, lethal complication of measles caused by particular mutants of measles virus (MeV that persist in the brain despite high levels of neutralizing antibodies. We addressed the hypothesis that antigenic drift is involved in the pathogenetic mechanism of SSPE by analyzing antigenic alterations in the MeV envelope hemagglutinin protein (MeV-H found in patients with SSPE in relation to major circulating MeV genotypes. To this aim, we obtained cDNA for the MeV-H gene from tissue taken at brain autopsy from 3 deceased persons with SSPE who had short (3-4 months, SMa79, average (3.5 years, SMa84, and long (18 years, SMa94 disease courses. Recombinant MeVs with a substituted MeV-H gene were generated by a reverse genetic system. Virus neutralization assays with a panel of anti-MeV-H murine monoclonal antibodies (mAbs or vaccine-immunized mouse anti-MeV-H polyclonal sera were performed to determine the antigenic relatedness. Functional and receptor-binding analysis of the SSPE MeV-H showed activity in a SLAM/nectin-4-dependent manner. Similar to our panel of wild-type viruses, our SSPE viruses showed an altered antigenic profile. Genotypes A, G3, and F (SSPE case SMa79 were the exception, with an intact antigenic structure. Genotypes D7 and F (SSPE SMa79 showed enhanced neutralization by mAbs targeting antigenic site IIa. Genotypes H1 and the recently reported D4.2 were the most antigenically altered genotypes. Epitope mapping of neutralizing mAbs BH015 and BH130 reveal a new antigenic site on MeV-H, which we designated Φ for its intermediate position between previously defined antigenic sites Ia and Ib. We conclude that SSPE-causing viruses show similar antigenic properties to currently circulating MeV genotypes. The absence of a direct correlation between antigenic changes and predisposition of a certain genotype to cause SSPE does not lend support to the proposed antigenic drift as a

Lethal interpersonal violence in the Middle Pleistocene.

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Nohemi Sala

Full Text Available Evidence of interpersonal violence has been documented previously in Pleistocene members of the genus Homo, but only very rarely has this been posited as the possible manner of death. Here we report the earliest evidence of lethal interpersonal violence in the hominin fossil record. Cranium 17 recovered from the Sima de los Huesos Middle Pleistocene site shows two clear perimortem depression fractures on the frontal bone, interpreted as being produced by two episodes of localized blunt force trauma. The type of injuries, their location, the strong similarity of the fractures in shape and size, and the different orientations and implied trajectories of the two fractures suggest they were produced with the same object in face-to-face interpersonal conflict. Given that either of the two traumatic events was likely lethal, the presence of multiple blows implies an intention to kill. This finding shows that the lethal interpersonal violence is an ancient human behavior and has important implications for the accumulation of bodies at the site, supporting an anthropic origin.

Lethal interpersonal violence in the Middle Pleistocene.

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Sala, Nohemi; Arsuaga, Juan Luis; Pantoja-Pérez, Ana; Pablos, Adrián; Martínez, Ignacio; Quam, Rolf M; Gómez-Olivencia, Asier; Bermúdez de Castro, José María; Carbonell, Eudald

2015-01-01

Evidence of interpersonal violence has been documented previously in Pleistocene members of the genus Homo, but only very rarely has this been posited as the possible manner of death. Here we report the earliest evidence of lethal interpersonal violence in the hominin fossil record. Cranium 17 recovered from the Sima de los Huesos Middle Pleistocene site shows two clear perimortem depression fractures on the frontal bone, interpreted as being produced by two episodes of localized blunt force trauma. The type of injuries, their location, the strong similarity of the fractures in shape and size, and the different orientations and implied trajectories of the two fractures suggest they were produced with the same object in face-to-face interpersonal conflict. Given that either of the two traumatic events was likely lethal, the presence of multiple blows implies an intention to kill. This finding shows that the lethal interpersonal violence is an ancient human behavior and has important implications for the accumulation of bodies at the site, supporting an anthropic origin.

The Effect of Total Cumulative Dose, Number of Treatment Cycles, Interval between Injections, and Length of Treatment on the Frequency of Occurrence of Antibodies to Botulinum Toxin Type A in the Treatment of Muscle Spasticity

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Bakheit, Abdel Magid O.; Liptrot, Anthea; Newton, Rachel; Pickett, Andrew M.

2012-01-01

A large cumulative dose of botulinum toxin type A (BoNT-A), frequent injections, a short interval between treatment cycles, and a long duration of treatment have all been suggested, but not confirmed, to be associated with a high incidence of neutralizing antibodies to the neurotoxin. The aim of this study was to investigate whether these…

A Quantitative Electrochemiluminescence Assay for Clostridium perfringens alpha toxin

National Research Council Canada - National Science Library

Merrill, Gerald A; Rivera, Victor R; Neal, Dwayne D; Young, Charles; Poli, Mark A

2006-01-01

.... Biotinylated antibodies to C. perfringens alpha toxin bound to streptavidin paramagnetic beads specifically immunoadsorbed soluble sample alpha toxin which subsequently selectively immunoadsorbed ruthenium (Ru...

Fate of Fusarium Toxins during Brewing.

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Habler, Katharina; Geissinger, Cajetan; Hofer, Katharina; Schüler, Jan; Moghari, Sarah; Hess, Michael; Gastl, Martina; Rychlik, Michael

2017-01-11

Some information is available about the fate of Fusarium toxins during the brewing process, but only little is known about the single processing steps in detail. In our study we produced beer from two different barley cultivars inoculated with three different Fusarium species, namely, Fusarium culmorum, Fusarium sporotrichioides, and Fusarium avenaceum, producing a wide range of mycotoxins such as type B trichothecenes, type A trichothecenes, and enniatins. By the use of multi-mycotoxin LC-MS/MS stable isotope dilution methods we were able to follow the fate of Fusarium toxins during the entire brewing process. In particular, the type B trichothecenes deoxynivalenol, 3-acetyldeoxynivalenol, and 15-acetyldeoxynivalenol showed similar behaviors. Between 35 and 52% of those toxins remained in the beer after filtration. The contents of the potentially hazardous deoxynivalenol-3-glucoside and the type A trichothecenes increased during mashing, but a rapid decrease of deoxynivalenol-3-glucoside content was found during the following steps of lautering and wort boiling. The concentration of enniatins greatly decreased with the discarding of spent grains or finally with the hot break. The results of our study show the retention of diverse Fusarium toxins during the brewing process and allow for assessing the food safety of beer regarding the monitored Fusarium mycotoxins.

Evolution of Bacillus thuringiensis Cry toxins insecticidal activity.

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Bravo, Alejandra; Gómez, Isabel; Porta, Helena; García-Gómez, Blanca Ines; Rodriguez-Almazan, Claudia; Pardo, Liliana; Soberón, Mario

2013-01-01

Insecticidal Cry proteins produced by Bacillus thuringiensis are use worldwide in transgenic crops for efficient pest control. Among the family of Cry toxins, the three domain Cry family is the better characterized regarding their natural evolution leading to a large number of Cry proteins with similar structure, mode of action but different insect specificity. Also, this group is the better characterized regarding the study of their mode of action and the molecular basis of insect specificity. In this review we discuss how Cry toxins have evolved insect specificity in nature and analyse several cases of improvement of Cry toxin action by genetic engineering, some of these examples are currently used in transgenic crops. We believe that the success in the improvement of insecticidal activity by genetic evolution of Cry toxins will depend on the knowledge of the rate-limiting steps of Cry toxicity in different insect pests, the mapping of the specificity binding regions in the Cry toxins, as well as the improvement of mutagenesis strategies and selection procedures. © 2012 The Authors. Microbial Biotechnology © 2012 Society for Applied Microbiology and Blackwell Publishing Ltd.

Treatment of Palatal Myoclonus with Botulinum Toxin Injection

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Mursalin M. Anis

2013-01-01

Full Text Available Palatal myoclonus is a rare cause of pulsatile tinnitus in patients presenting to the otolaryngology office. Rhythmic involuntary contractions of the palatal muscles produce the pulsatile tinnitus in these patients. Treatment of this benign but distressing condition with anxiolytics, anticonvulsants, and surgery has been largely unsuccessful. A few investigators have obtained promising results with botulinum toxin injection into the palatal muscles. We present a patient with palatal myoclonus who failed conservative treatment with anxiolytics. Unilateral injection of botulinum toxin into her tensor veli palatini muscle under electromyographic guidance resolved pulsatile tinnitus in her ipsilateral ear and unmasked pulsatile tinnitus in the contralateral ear. A novel method of following transient postinjection symptoms using a diary is presented in this study. Botulinum toxin dose must be titrated to achieve optimal results in each individual patient, analogous to titrations done for spasmodic dysphonia. Knowledge of the temporal onset of postinjection side effects and symptomatic relief may aid physicians in dose titration and surveillance. We present suggestions on titrating the botulinum toxin dose to optimal levels. A review of the literature on the use of botulinum toxin for palatal myoclonus and some common complications are discussed.

Treatment of Gastrointestinal Sphincters Spasms with Botulinum Toxin A

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Giuseppe Brisinda

2015-05-01

Full Text Available Botulinum toxin A inhibits neuromuscular transmission. It has become a drug with many indications. The range of clinical applications has grown to encompass several neurological and non-neurological conditions. One of the most recent achievements in the field is the observation that botulinum toxin A provides benefit in diseases of the gastrointestinal tract. Although toxin blocks cholinergic nerve endings in the autonomic nervous system, it has also been shown that it does not block non-adrenergic non-cholinergic responses mediated by nitric oxide. This has promoted further interest in using botulinum toxin A as a treatment for overactive smooth muscles and sphincters. The introduction of this therapy has made the treatment of several clinical conditions easier, in the outpatient setting, at a lower cost and without permanent complications. This review presents current data on the use of botulinum toxin A in the treatment of pathological conditions of the gastrointestinal tract.

Cosmetic Effect of Botulinum Toxin In Focal Hyperhydrosis

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Jain S

2005-01-01

Full Text Available Hyperhydrosis of axillae, palm and sole is not a very uncommon problem. It leads to great embarrassment and considerable emotional stress to the individuals. Botulinum toxins prevent the release of acetylcholine at nerve terminals, therefore, reduces sweat secretion. Six patients of axillary and 4 patients of palmer and planter hyperhydrosis were treated with botulinum toxin. All patients experienced relatively satisfactory reduction of hyperhydrosis for period ranging between 4-7 months. No adverse effects were observed. Botulinum toxin therefore can be considered as an effective treatment in focal hyperhydrosis.

The resurgence of botulinum toxin injection for strabismus in children.

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Mahan, Marielle; Engel, J Mark

2017-09-01

The present review discusses recent advances in the use of botulinum toxin for the management of strabismus in children. Botulinum toxin injection produces similar results compared to surgery for certain subtypes of strabismus, especially acute onset esotropia. It may be more effective in many subtypes of esotropia where surgery has been less reliable, including partially accommodative esotropia, esotropia associated with cerebral palsy, and thyroid eye disease. Small retrospective studies have demonstrated the efficacy of botulinum toxin in the treatment of many types of pediatric strabismus, providing some guidance for clinicians to determine which patients would benefit most from this intervention. Although administration of botulinum toxin is generally accepted as a reasonable option in select cases, many strabismus surgeons have not fully embraced the treatment, in part because of perceived disadvantages compared to surgery and difficulty in identifying subsets with the highest potential for therapeutic success. A recent study compared the administration of botulinum toxin in children with acute-onset esotropia to surgical correction and found botulinum toxin had a statistically equal success rate, but with the advantage of significantly less time under general anesthesia. In addition, botulinum toxin has been recently tried in patients with partially accommodative esotropia, esotropia associated with cerebral palsy, cyclic esotropia, and in patients with thyroid eye disease. The present review will discuss current clinical recommendations based on recent studies on the use of botulinum toxin in children with strabismus.

General synthesis of β-alanine-containing spider polyamine toxins and discovery of nephila polyamine toxins 1 and 8 as highly potent inhibitors of ionotropic glutamate receptors

DEFF Research Database (Denmark)

Lucas, Simon; Poulsen, Mette H; Nørager, Niels G

2012-01-01

Certain spiders contain large pools of polyamine toxins, which are putative pharmacological tools awaiting further discovery. Here we present a general synthesis strategy for this class of toxins and prepare five structurally varied polyamine toxins. Electrophysiological testing at three ionotrop...

Effect of Gating Modifier Toxins on Membrane Thickness: Implications for Toxin Effect on Gramicidin and Mechanosensitive Channels

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Shin-Ho Chung

2013-02-01

Full Text Available Various gating modifier toxins partition into membranes and interfere with the gating mechanisms of biological ion channels. For example, GsMTx4 potentiates gramicidin and several bacterial mechanosensitive channels whose gating kinetics are sensitive to mechanical properties of the membrane, whereas binding of HpTx2 shifts the voltage-activity curve of the voltage-gated potassium channel Kv4.2 to the right. The detailed process by which the toxin partitions into membranes has been difficult to probe using molecular dynamics due to the limited time scale accessible. Here we develop a protocol that allows the spontaneous assembly of a polypeptide toxin into membranes in atomistic molecular dynamics simulations of tens of nanoseconds. The protocol is applied to GsMTx4 and HpTx2. Both toxins, released in water at the start of the simulation, spontaneously bind into the lipid bilayer within 50 ns, with their hydrophobic patch penetrated into the bilayer beyond the phosphate groups of the lipids. It is found that the bilayer is about 2 Å thinner upon the binding of a GsMTx4 monomer. Such a thinning effect of GsMTx4 on membranes may explain its potentiation effect on gramicidin and mechanosensitive channels.

Potent neutralization of influenza A virus by a single-domain antibody blocking M2 ion channel protein.

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Guowei Wei

Full Text Available Influenza A virus poses serious health threat to humans. Neutralizing antibodies against the highly conserved M2 ion channel is thought to offer broad protection against influenza A viruses. Here, we screened synthetic Camel single-domain antibody (VHH libraries against native M2 ion channel protein. One of the isolated VHHs, M2-7A, specifically bound to M2-expressed cell membrane as well as influenza A virion, inhibited replication of both amantadine-sensitive and resistant influenza A viruses in vitro, and protected mice from a lethal influenza virus challenge. Moreover, M2-7A showed blocking activity for proton influx through M2 ion channel. These pieces of evidence collectively demonstrate for the first time that a neutralizing antibody against M2 with broad specificity is achievable, and M2-7A may have potential for cross protection against a number of variants and subtypes of influenza A viruses.

Potentiometric chemical sensors for the detection of paralytic shellfish toxins.

Science.gov (United States)

Ferreira, Nádia S; Cruz, Marco G N; Gomes, Maria Teresa S R; Rudnitskaya, Alisa

2018-05-01

Potentiometric chemical sensors for the detection of paralytic shellfish toxins have been developed. Four toxins typically encountered in Portuguese waters, namely saxitoxin, decarbamoyl saxitoxin, gonyautoxin GTX5 and C1&C2, were selected for the study. A series of miniaturized sensors with solid inner contact and plasticized polyvinylchloride membranes containing ionophores, nine compositions in total, were prepared and their characteristics evaluated. Sensors displayed cross-sensitivity to four studied toxins, i.e. response to several toxins together with low selectivity. High selectivity towards paralytic shellfish toxins was observed in the presence of inorganic cations with selectivity coefficients ranging from 0.04 to 0.001 for Na + and K + and 3.6*10 -4 to 3.4*10 -5 for Ca 2+ . Detection limits were in the range from 0.25 to 0.9 μmolL -1 for saxitoxin and decarbamoyl saxitoxin, and from 0.08 to 1.8 μmolL -1 for GTX5 and C1&C2, which allows toxin detection at the concentration levels corresponding to the legal limits. Characteristics of the developed sensors allow their use in the electronic tongue multisensor system for simultaneous quantification of paralytic shellfish toxins. Copyright © 2018 Elsevier B.V. All rights reserved.

Transporting Patients with Lethal Contagious Infections

National Research Council Canada - National Science Library

Swartz, Colleen

2002-01-01

.... The AIT is a unique military medical team capable of worldwide air evacuation and management of a limited number of patients who are potentially exposed to known and unknown lethal communicable...

Milling technological experiments to reduce Fusarium toxin contamination in wheat

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Véha A.

2015-01-01

Full Text Available We examine 4 different DON-toxin-containing (0.74 - 1.15 - 1.19 - 2.14 mg/kg winter wheat samples: they were debranned and undebranned, and we investigated the flour’s and the by-products’ (coarse, fine bran toxin content changes. SATAKE lab-debranner was used for debranning and BRABENDER lab-mill for the milling process. Without debranning, two sample flours were above the DON toxin limit (0.75 mg/kg, which are waste. By minimum debranning (and minimum debranning mass loss; 6-8%, our experience with whole flour is that the multi-stage debranning measurement significantly reduces the content of the flour’s DON toxin, while the milling by-products, only after careful consideration and DON toxin measurements, may be produced for public consumption and for feeding.

A bacterial cocaine esterase protects against cocaine-induced epileptogenic activity and lethality.

Science.gov (United States)

Jutkiewicz, Emily M; Baladi, Michelle G; Cooper, Ziva D; Narasimhan, Diwahar; Sunahara, Roger K; Woods, James H

2009-09-01

Cocaine toxicity results in cardiovascular complications, seizures, and death and accounts for approximately 20% of drug-related emergency department visits every year. Presently, there are no treatments to eliminate the toxic effects of cocaine. The present study hypothesizes that a bacterial cocaine esterase with high catalytic efficiency would provide rapid and robust protection from cocaine-induced convulsions, epileptogenic activity, and lethality. Cocaine-induced paroxysmal activity and convulsions were evaluated in rats surgically implanted with radiotelemetry devices (N=6 per treatment group). Cocaine esterase was administered 1 minute after a lethal dose of cocaine or after cocaine-induced convulsions to determine the ability of the enzyme to prevent or reverse, respectively, the effects of cocaine. The cocaine esterase prevented all cocaine-induced electroencephalographic changes and lethality. This effect was specific for cocaine because the esterase did not prevent convulsions and death induced by a cocaine analog, (-)-2beta-carbomethoxy-3beta-phenyltropane. The esterase prevented lethality even after cocaine-induced convulsions occurred. In contrast, the short-acting benzodiazepine, midazolam, prevented cocaine-induced convulsions but not the lethal effects of cocaine. The data showed that cocaine esterase successfully degraded circulating cocaine to prevent lethality and that cocaine-induced convulsions alone are not responsible for the lethal effects of cocaine in this model. Therefore, further investigation into the use of cocaine esterase for treating cocaine overdose and its toxic effects is warranted.

Dominant-lethal mutations and heritable translocations in mice

Energy Technology Data Exchange (ETDEWEB)

Generoso, W.M.

1983-01-01

Chromosome aberrations are a major component of radiation or chemically induced genetic damage in mammalian germ cells. The types of aberration produced are dependent upon the mutagen used and the germ-cell stage treated. For example, in male meiotic and postmeiotic germ cells certain alkylating chemicals induce both dominant-lethal mutations and heritable translocations while others induce primarily dominant-lethal mutations. Production of these two endpoints appears to be determined by the stability of alkylation products with the chromosomes. If the reaction products are intact in the male chromosomes at the time of sperm entry, they may be repaired in fertilized eggs. If repair is not effected and the alkylation products persist to the time of pronuclear chromosome replication, they lead to chromatid-type aberrations and eventually to dominant-lethality. The production of heritable translocations, on the other hand, requires a transformation of unstable alkylation products into suitable intermediate lesions. The process by which these lesions are converted into chromosome exchange within the male genome takes place after sperm enters the egg but prior to the time of pronuclear chromosome replication (i.e., chromosome-type). Thus, dominant-lethal mutations result from both chromatid- and chromosome-type aberrations while heritable translocations result primarily from the latter type. DNA target sites associated with the production of these two endpoints are discussed.

Dominant-lethal mutations and heritable translocations in mice

International Nuclear Information System (INIS)

Generoso, W.M.

1983-01-01

Chromosome aberrations are a major component of radiation or chemically induced genetic damage in mammalian germ cells. The types of aberration produced are dependent upon the mutagen used and the germ-cell stage treated. For example, in male meiotic and postmeiotic germ cells certain alkylating chemicals induce both dominant-lethal mutations and heritable translocations while others induce primarily dominant-lethal mutations. Production of these two endpoints appears to be determined by the stability of alkylation products with the chromosomes. If the reaction products are intact in the male chromosomes at the time of sperm entry, they may be repaired in fertilized eggs. If repair is not effected and the alkylation products persist to the time of pronuclear chromosome replication, they lead to chromatid-type aberrations and eventually to dominant-lethality. The production of heritable translocations, on the other hand, requires a transformation of unstable alkylation products into suitable intermediate lesions. The process by which these lesions are converted into chromosome exchange within the male genome takes place after sperm enters the egg but prior to the time of pronuclear chromosome replication (i.e., chromosome-type). Thus, dominant-lethal mutations result from both chromatid- and chromosome-type aberrations while heritable translocations result primarily from the latter type. DNA target sites associated with the production of these two endpoints are discussed

Staphylococcus aureus α-toxin modulates skin host response to viral infection.

Science.gov (United States)

Bin, Lianghua; Kim, Byung Eui; Brauweiler, Anne; Goleva, Elena; Streib, Joanne; Ji, Yinduo; Schlievert, Patrick M; Leung, Donald Y M

2012-09-01

Patients with atopic dermatitis (AD) with a history of eczema herpeticum have increased staphylococcal colonization and infections. However, whether Staphylococcus aureus alters the outcome of skin viral infection has not been determined. We investigated whether S aureus toxins modulated host response to herpes simplex virus (HSV) 1 and vaccinia virus (VV) infections in normal human keratinocytes (NHKs) and in murine infection models. NHKs were treated with S aureus toxins before incubation of viruses. BALB/c mice were inoculated with S aureus 2 days before VV scarification. Viral loads of HSV-1 and VV were evaluated by using real-time PCR, a viral plaque-forming assay, and immunofluorescence staining. Small interfering RNA duplexes were used to knockdown the gene expression of the cellular receptor of α-toxin, a disintegrin and metalloprotease 10 (ADAM10). ADAM10 protein and α-toxin heptamers were detected by using Western blot assays. We demonstrate that sublytic staphylococcal α-toxin increases viral loads of HSV-1 and VV in NHKs. Furthermore, we demonstrate in vivo that the VV load is significantly greater (P skin inoculated with an α-toxin-producing S aureus strain compared with murine skin inoculated with the isogenic α-toxin-deleted strain. The viral enhancing effect of α-toxin is mediated by ADAM10 and is associated with its pore-forming property. Moreover, we demonstrate that α-toxin promotes viral entry in NHKs. The current study introduces the novel concept that staphylococcal α-toxin promotes viral skin infection and provides a mechanism by which S aureus infection might predispose the host toward disseminated viral infections. Copyright © 2012 American Academy of Allergy, Asthma & Immunology. Published by Mosby, Inc. All rights reserved.

Quantitative determination of biological activity of botulinum toxins utilizing compound muscle action potentials (CMAP), and comparison of neuromuscular transmission blockage and muscle flaccidity among toxins.

Science.gov (United States)

Torii, Yasushi; Goto, Yoshitaka; Takahashi, Motohide; Ishida, Setsuji; Harakawa, Tetsuhiro; Sakamoto, Takashi; Kaji, Ryuji; Kozaki, Shunji; Ginnaga, Akihiro

2010-01-01

The biological activity of various types of botulinum toxin has been evaluated using the mouse intraperitoneal LD(50) test (ip LD(50)). This method requires a large number of mice to precisely determine toxin activity, and so has posed a problem with regard to animal welfare. We have used a direct measure of neuromuscular transmission, the compound muscle action potential (CMAP), to evaluate the effect of different types of botulinum neurotoxin (NTX), and we compared the effects of these toxins to evaluate muscle relaxation by employing the digit abduction scoring (DAS) assay. This method can be used to measure a broad range of toxin activities the day after administration. Types A, C, C/D, and E NTX reduced the CMAP amplitude one day after administration at below 1 ip LD(50), an effect that cannot be detected using the mouse ip LD(50) assay. The method is useful not only for measuring toxin activity, but also for evaluating the characteristics of different types of NTX. The rat CMAP test is straightforward, highly reproducible, and can directly determine the efficacy of toxin preparations through their inhibition of neuromuscular transmission. Thus, this method may be suitable for pharmacology studies and the quality control of toxin preparations. Copyright 2009 Elsevier Ltd. All rights reserved.

Prediction of Toxin Genes from Chinese Yellow Catfish Based on Transcriptomic and Proteomic Sequencing

Directory of Open Access Journals (Sweden)

Bing Xie

2016-04-01

Full Text Available Fish venom remains a virtually untapped resource. There are so few fish toxin sequences for reference, which increases the difficulty to study toxins from venomous fish and to develop efficient and fast methods to dig out toxin genes or proteins. Here, we utilized Chinese yellow catfish (Pelteobagrus fulvidraco as our research object, since it is a representative species in Siluriformes with its venom glands embedded in the pectoral and dorsal fins. In this study, we set up an in-house toxin database and a novel toxin-discovering protocol to dig out precise toxin genes by combination of transcriptomic and proteomic sequencing. Finally, we obtained 15 putative toxin proteins distributed in five groups, namely Veficolin, Ink toxin, Adamalysin, Za2G and CRISP toxin. It seems that we have developed a novel bioinformatics method, through which we could identify toxin proteins with high confidence. Meanwhile, these toxins can also be useful for comparative studies in other fish and development of potential drugs.

Pufferfish mortality associated with novel polar marine toxins in Hawaii

Science.gov (United States)

Work, Thierry M.; Moeller, Perer D. R.; Beauchesne, Kevin R.; Dagenais, Julie; Breeden, Renee; Rameyer, Robert; Walsh, Willliam A.; Abecassis, Melanie; Kobayashi, Donald R.; Conway, Carla M.; Winton, James

2017-01-01

Fish die-offs are important signals in tropical marine ecosystems. In 2010, a mass mortality of pufferfish in Hawaii (USA) was dominated by Arothron hispidus showing aberrant neurological behaviors. Using pathology, toxinology, and field surveys, we implicated a series of novel, polar, marine toxins as a likely cause of this mass mortality. Our findings are striking in that (1) a marine toxin was associated with a kill of a fish species that is itself toxic; (2) we provide a plausible mechanism to explain clinical signs of affected fish; and (3) this epizootic likely depleted puffer populations. Whilst our data are compelling, we did not synthesize the toxin de novo, and we were unable to categorically prove that the polar toxins caused mortality or that they were metabolites of an undefined parent compound. However, our approach does provide a template for marine fish kill investigations associated with marine toxins and inherent limitations of existing methods. Our study also highlights the need for more rapid and cost-effective tools to identify new marine toxins, particularly small, highly polar molecules.

ADP-ribosylation by cholera toxin: functional analysis of a cellular system that stimulates the enzymic activity of cholera toxin fragment A1

International Nuclear Information System (INIS)

Gill, D.M.; Coburn, J.

1987-01-01

The authors have clarified relationships between cholera toxin, cholera toxin substrates, a membrane protein S that is required for toxin activity, and a soluble protein CF that is needed for the function of S. The toxin has little intrinsic ability to catalyze ADP-ribosylations unless it encounters the active form of the S protein, which is S liganded to GTP or to a GTP analogue. In the presence of CF, S x GTP forms readily, though reversibly, but a more permanent active species, S-guanosine 5'-O-(3-thiotriphosphate) (S x GTPγS), forms over a period of 10-15 min at 37 0 C. Both guanosine 5'-O-(2-thiodiphosphate) and GTP block this quasi-permanent activation. Some S x GTPγS forms in membranes that are exposed to CF alone and then to GTPγS, with a wash in between, and it is possible that CF facilitates a G nucleotide exchange. S x GTPγS dissolved by nonionic detergents persists in solution and can be used to support the ADP-ribosylation of nucleotide-free substrates. In this circumstance, added guanyl nucleotides have no further effect. This active form of S is unstable, especially when heated, but the thermal inactivation above 45 0 C is decreased by GTPγS. Active S is required equally for the ADP-ribosylation of all of cholera toxin's protein substrates, regardless of whether they bind GTP or not. They suggest that active S interacts directly with the enzymic A 1 fragments of cholera toxin and not with any toxin substrate. The activation and activity of S are independent of the state, or even the presence, of adenylate cyclase and seem to be involved with the cyclase system only via cholera toxin. S is apparently not related by function to certain other GTP binding proteins, including p21/sup ras/, and appears to be a new GTP binding protein whose physiologic role remains to be identified

A lethal ovitrap-based mass trapping scheme for dengue control in Australia: I. Public acceptability and performance of lethal ovitraps.

Science.gov (United States)

Ritchie, S A; Rapley, L P; Williams, C; Johnson, P H; Larkman, M; Silcock, R M; Long, S A; Russell, R C

2009-12-01

We report on the first field evaluation of the public acceptability and performance of two types of lethal ovitrap (LO) in three separate trials in Cairns, Australia. Health workers were able to set standard lethal ovitraps (SLOs) in 75 and 71% of premise yards in the wet and dry season, respectively, and biodegradable lethal ovitraps (BLOs) in 93% of yards. Public acceptance, measured as retention of traps by residents, was high for both trap types, with porous (grass, soil and mulch) versus solid (tiles, concrete, wood and stone) substrates. The SLOs and the BLOs were readily acceptable to ovipositing Aedes aegypti L. (Diptera: Culicidae); the mean number of eggs/trap was 6 and 15, for the dry season and wet season SLO trial, respectively, and 15 for the BLO wet season trial. Indeed, 84-94% of premise yards had egg positive SLOs or BLOs. A high percentage of both wet and dry season SLOs (29 and 70%, respectively) and BLOs (62%) that were dry after 4 weeks were egg positive, indicating the traps had functioned. Lethal strips from SLOs and BLOs that had been exposed for 4 weeks killed 83 and 74%, respectively, of gravid Ae. aegypti in laboratory assays. These results indicate that mass trapping schemes using SLOs and BLOs are not rejected by the public and effectively target gravid Ae. aegypti. The impact of the interventions on mosquito populations is described in a companion paper.

Structural constraints-based evaluation of immunogenic avirulent toxins from Clostridium botulinum C2 and C3 toxins as subunit vaccines.

Science.gov (United States)

Prisilla, A; Prathiviraj, R; Sasikala, R; Chellapandi, P

2016-10-01

Clostridium botulinum (group-III) is an anaerobic bacterium producing C2 and C3 toxins in addition to botulinum neurotoxins in avian and mammalian cells. C2 and C3 toxins are members of bacterial ADP-ribosyltransferase superfamily, which modify the eukaryotic cell surface proteins by ADP-ribosylation reaction. Herein, the mutant proteins with lack of catalytic and pore forming function derived from C2 (C2I and C2II) and C3 toxins were computationally evaluated to understand their structure-function integrity. We have chosen many structural constraints including local structural environment, folding process, backbone conformation, conformational dynamic sub-space, NAD-binding specificity and antigenic determinants for screening of suitable avirulent toxins. A total of 20 avirulent mutants were identified out of 23 mutants, which were experimentally produced by site-directed mutagenesis. No changes in secondary structural elements in particular to α-helices and β-sheets and also in fold rate of all-β classes. Structural stability was maintained by reordered hydrophobic and hydrogen bonding patterns. Molecular dynamic studies suggested that coupled mutations may restrain the binding affinity to NAD(+) or protein substrate upon structural destabilization. Avirulent toxins of this study have stable energetic backbone conformation with a common blue print of folding process. Molecular docking studies revealed that avirulent mutants formed more favorable hydrogen bonding with the side-chain of amino acids near to conserved NAD-binding core, despite of restraining NAD-binding specificity. Thus, structural constraints in the avirulent toxins would determine their immunogenic nature for the prioritization of protein-based subunit vaccine/immunogens to avian and veterinary animals infected with C. botulinum. Copyright © 2016 Elsevier B.V. All rights reserved.

Lethal mechanisms in gastric volvulus.

Science.gov (United States)

Omond, Kimberley J; Byard, Roger W

2017-01-01

A 55-year-old wheelchair-bound woman with severe cerebral palsy was found at autopsy to have marked distention of the stomach due to a volvulus. The stomach was viable, and filled with air and fluid and had pushed the left dome of the diaphragm upwards causing marked compression of the left lung with a mediastinal shift to the right (including the heart). There was no evidence of gastric perforation, ischaemic necrosis or peritonitis. Removal of the organ block revealed marked kyphoscoliosis. Histology confirmed the viability of the stomach and biochemistry showed no dehydration. Death in cases of acute gastric volvulus usually occurs because of compromise of the gastric blood supply resulting in ischaemic necrosis with distention from swallowed air and fluid resulting in perforation with lethal peritonitis. Hypovolaemic shock may also occur. However, the current case demonstrates an alternative lethal mechanism, that of respiratory compromise due to marked thoracic organ compression.

Toxin-mediated effects on the innate mucosal defenses: implications for enteric vaccines

DEFF Research Database (Denmark)

Glenn, Gregory M; Francis, David H; Danielsen, E Michael

2009-01-01

mucosal barrier as a key step in enteric pathogen survival. We review key observations relevant to the roles of LT and cholera toxin in protective immunity and the effects of these toxins on innate mucosal defenses. We suggest either that toxin-mediated fluid secretion mechanically disrupts the mucus...... layer or that toxins interfere with innate mucosal defenses by other means. Such a breach gives pathogens access to the enterocyte, leading to binding and pathogenicity by enterotoxigenic E. coli (ETEC) and other organisms. Given the common exposure to LT(+) ETEC by humans visiting or residing...... unexpectedly broad protective effects against LT(+) ETEC and mixed infections when using a toxin-based enteric vaccine. If toxins truly exert barrier-disruptive effects as a key step in pathogenesis, then a return to classic toxin-based vaccine strategies for enteric disease is warranted and can be expected...