Animal Toxins: state of the art: perspectives in health and biotecnology

Apresenta, pela primeira vez em um único volume, o resultado de diversas pesquisas com diferentes perspectivas sobre as reais possibilidades do uso de venenos e de toxinas de origem animal na indústria biotecnológica. Animal Toxins é composto por 39 artigos, em inglês, assinados por especialistas renomados de várias nacionalidades. O estado da arte em compostos obtidos a partir de venenos de animais marinhos, aranhas e escorpiões, lagartas, serpentes, entre outros, são o foco desta publicação que visa a atender cientistas, estudantes e pesquisadores da universidade, da indústria farmacêutica e de biotecnologia interessados em toxicologia. Os venenos e toxinas de origem animal foram selecionados ao longo de milhões de anos de evolução para atuarem de forma rápida e eficaz no organismo da vítima, o que resultou em um repertorio maciço de moléculas capazes de se ligarem a alvos específicos. A possibilidade de utilização dessas toxinas em processos biotecnológicos fez com que esses venenos e toxinas sejam considerados como uma das mais promissoras fontes de compostos bioativos naturais.

Evaluation of experimental conditions for quantification of LT produced by human derived enterotoxigenic Escherichia coli strains

The heat-labile toxin (LT) is a key virulence-associated factor associated with the non-invasive secretory diarrhea caused by enterotoxigenic Escherichia coli (ETEC) strains either in humans or domestic animals. Several LT detection methods have been reported but quantification of the toxin produced by wild-type ETEC strains is usually performed by the GM1 ganglyoside enzyme-linked immunosorbent assay (GM1 ELISA). In this study we conducted the optimization of an alternative LT-quantification method, the antibody-capture ELISA (cELISA). Detailed analysis of the appropriate dilutions of capture and detecting LT-specific antibodies significantly improved the sensitivity of the method. Additionally, testing of different LT extraction techniques indicated that sonic disruption of the bacterial cells enhanced LT recovery yields, in contrast to the usual procedure based on addition of polymyxin B to the culture medium as well as extraction methods based on chloroform or Triton X-100. Moreover, the present data indicate that performance of the LT extraction method based on polymyxin B treatment can vary among wild ETEC strains.

A toxina termo-lábil (LT) é um fator de virulência associado à diarréia secretora não invasiva causada por linhagens de Escherichia coli enterotoxigênica (ETEC) em humanos ou animais domésticos. Diversos métodos de detecção de LT foram descritos na literatura, no entanto, a quantificação da toxina produzida por linhagens selvagens de ETEC é geralmente realizada por ensaio imunoenzimático com o gangliosídeo GM-1 (GM-1 ELISA). Neste estudo, conduzimos uma otimização experimental de um método alternativo de quantificação de LT, o ELISA de captura (cELISA). Análise detalhada de diluições apropriadas dos anticorpos LT específicos de captura e detecção melhorou significantemente a sensibilidade do método. Em adição, testes com diferentes técnicas de extração de LT indicaram que a ruptura das células por ultra-som, mas não o tratamento com polimixina B, clorofórmio ou Triton X-100, aumentou o rendimento da recuperação de LT. Além disto, os dados apresentados demonstram que o desempenho do método de extração de LT baseado no tratamento com polimixina B pode variar entre linhagens selvagens de ETEC.

Purification and characterization of toxins from wheat isolates of Drechslera tritici-repentis, Bipolaris bicolor, and Bipolaris sorokiniana

Low molecular weight metabolites produced by Bipolaris bicolor, Bipolaris sorokiniana, and Drechslera tritici-repentis are considered to be toxins that facilitate disease in wheat cultivars. Several such toxins were isolated from these fungi. Electrophoresis demonstrated bands of proteins that reduce shoot inhibition in susceptible plants but not in resistant plants. Chlorophyll content was reduced during the first 10 hours of light in the susceptible plants and after 18 hours in the resistant plants. The enzyme beta-1,3-glucanase increased in the resistant plants after treatment with toxins, but in the susceptible plants this enzyme decreased compared to the control. This suggests that the toxin was a protein and the susceptible plants needed other mechanisms for induce resistance.

Purification and characterization of a shiga-like toxin from escherichia coli strain 0157 of human origin implicated in haemolytic uremic syndrome

A Shiga-like toxin [SLT] from enterohemorrhagic Escherichia coli strain 0157 isolated from the stools of patients with haemolytic uremic syndrome [HUS] was produced, partially purified and the biological properties have been characterized. The production of the toxin was done on Tryptone Soy Broth containing mitomycin C. The purification scheme involved polymyxin beta extraction of the toxin from bacterial cell, differential [NH[4]]2 SO[4]. Precipitation and desalting by passage through a Sephadex G50 column chromatography equilibrated with 10 mM. Tris hydrochloride [pH 7.6]. The purification scheme resulted in high recovery of SLT activity in comparison to crude culture supernatant and the recovered cytotoxicity to Vero cells were 4 x 10[4] CD[50]/mg protein for crude culture supernatant, 1.8 x 10[5] CD[50]/mg for polymyxin B extract and 5 x 10[5] for the material obtained by [NH[4][2] SO[4]. fractionation and gel filtration. The partially purified SLT or Verotoxin was [I] Paralytic-lethal to mice when injiected intraperitoneally and the 50% lethal dose was [16.000] verotoxin units [ii] enterotoxic to rabbit ileal loop, at a very high dose [256, 000] verotoxoin unit, The purified toxin preparation was Partially homogenous as judged by Sodium dodecyl sulphate-polyacrylamide gel electrophoresis [SDS-PAGE]. The SLT preparation, under non reducing condition, exhibited 2 bands on SDS-PAGE corresponding to a molecolar mass of 33,000 and 7,500 kb together with other protein band, These 2 protein bands were consistent with the expected MWs of A and B subunits of SLT respectively

In vitro activity of the killer toxin from the yeast hansenula anomala against some different microorganisms

The killer toxin of Hansenula anomala showed killer activity toward some different microrganisms. The killer toxin produced by this species was purified. This toxin inhibited completely the growth of 2 species of yeasts,. Candida albicans, and CKefyr; three species of dermatophytes, Microsporum gypseum, M. Canis and Trichophyton mentagrophytes; and 2 species of moulds Fusarium solani and Aspergillus terreus tested at concentrations 100 ul/ml or less. The minimum inhibitory concentration [MIC] of the killer toxin against Candida guilliermondii and C. parapsilosis tested was 20 ul/ml or more. [MIC] of the killer toxin against, M. audoiunii, Sepedomium chrysospermun and Penicillium Coryophilum tested was 60 ul/ml or more. Finally the [MIC] of this killer toxin against the following bacterial strains: Proteus Vulgaris. Staphylococcus aureus, Salmonella typh, E. coli and Rhizobium legumnosarum tested was 25 ul/ml or more. But the [MIC] of this toxin against the two species of: Bacillus subtilus and Pseudomonas areuginosa was 50 ul/ml or more. In this paper we found that the purified H. anomala killer toxin used for the determination the [MIC] of a wide range of yeasts, moulds and bacteria including a number of important species

Isolation, characterization and biological activities of a toxin from Bacillus megaterium (B-23).

Fungitoxic substance was isolated from the culture filtrate of B. megaterium (B-23). Age of culture and pH of medium influence the fungitoxicity of its culture filtrate. Partially purified toxin was thermolabile, non-dialysable, ethyl acetate soluble, vanillin-sulphuric acid positive and effective within a range of pH 5-9. It exhibited maximum UV absorption at 224 nm. Its melting point was 242 degrees C. The efficacy of this compound was tested on 4 jute parasites namely, C. corchori, C. gloeosporioides, M. roridum and A. citri, of which M. roridum and C. corchori were least and most sensitive to the toxin respectively.

Lysis of red blood cells by extracts from benthic dinoflagellates

Samples of the cultured benthic dinoflagellates Gambierdiscus toxicus and Ostreopsis lenticularis, both isolated from a shalow back reef habitat in soutwestern Puerto Rico, were estracted in methanol, dried and resuspended in distilled water. After centrifugation, aliquots of the supernatant, or dilutions thereof, were added to suspensions of washed human and mouse red blood cells and incuated at different temperatures for different time periods. Further spectrophotometrical examinations of the samples showed a hemolytic activity aginst mouse and human red blood cells. The hemolytic activity of G. toxicus extract was 3 to 4 times greater than that of O. lenticularis and was less temperature-dependent. Such findings suggest that these two dinoflagellates produce chemically different hemolysins

Mecanismo de acción de miotoxinas aisladas de venenos de serpientes / Mechanism of action of myotoxins isolated from snake venoms

Desde los puntos de vista bioquímico y farmacológico, las miotoxinas aisladas de venenos de serpientes se ubican en cuatro grupos: (1) Fosfolipasas A miotóxicas, (2) miotoxinas básicas de bajo peso molecular, (3) cardiotoxinas de venenos elapídeos y (4) miotoxinas hemorrágicas. Las fosfolipasas miotóxicas notexinam taipoxina, crotoxina y miotoxina de Bothrops asper afectan inicialmente la integridad de la membrana plasmática, induciéndose un influjo de calcio que culmina con la muerte celular. Las miotoxinas básicas de bajo peso molecular crotamina y miotoxina a actúan específicamente en los canales de sodio del sarcolema, induciendo un influjo de sodio que trae como consecuencia despolarización y contracción muscular y vacuolización del retículo sarcoplásmico. Las cardiotoxinas son polipéptidos básicos capaces de desorganizar la estructura de las membranas, siendo su acción miotóxica una consecuencia de la alteración drástica que las mismas inducen en el sarcolema del músculo esquelético. Finalmente, dos componentes hemorrágicos (toxina hemorrágica b y viriditoxina) poseen actividad miotóxica, habiendose sugerido que este efecto es una consecuencia de la isquemia tisular resultante de la acción hemorrágica de estos componentes