Aedes aegypti (L.), the main vector of dengue fever in Brazil, has been controlled with the use of massive chemical products, contributing to the development of resistance and decreasing the insect control efficiency. The control of dipterans with bioinsecticides based on Bacillus thuringiensis has been satisfactory, due to the production of insecticidal proteins denominated Cry (crystal), Cyt (cytolytic) toxins and Chi (chitinase), and to the synergistic effects among them. The present work aimed to select B. thuringiensis isolates efficient against A. aegypti larvae. A bacterial collection containing 1,073 isolates of B. thuringiensis, obtained from different locations of Brazilian territory, had the DNA isolated and submitted to PCR amplifications using specific primers for cry4Aa, cry4Ba, cry11Aa, cry11Ba, cyt1Aa, cyt1Ab, cyt2Aa and chi genes. For the LC50 and LC90 determination, the entomopathogenic isolates were evaluated by selective and quantitative bioassays. Only 45 isolates (4.2%) presented amplicons for the cry and cyt genes. The chi gene sequence was detected in 25 (54.3%) of those isolates. From the 45 isolates submitted to the selective bioassays, 13 caused 100% mortality of A. aegypti larvae. The identification of cry, cyt and chi genes of B. thuringiensis and the toxicity analysis on A. aegypti led to the selection of a set of isolates that have the potential to be used in the formulation of new bioinsecticides.

O desenvolvimento da produção e uso do Bacillus thuringiensis no Brasil em escala comercial enfrenta certas dificuldades, entre elas o estabelecimento de metodologias para a quantificação de produtos tóxicos a serem comercializados. Atualmente, a quantidade de toxinas é expressa como porcentagem do total de proteínas presentes em amostras em consideração. Tal metodologia, entretanto, não mede a quantidade real de uma determinada proteína presente em um produto qualquer, além do fato de diferentes linhagens bacterianas possuírem diferentes genes codificadores para endotoxinas e mesmo para b-toxina. Desde que os diferentes tipos de toxinas apresentam diferentes características antigências, este trabalho tem como objetivo a utilização de técnicas imunológicas para quantificar específicamente o conteúdo de proteína cristal presente em diferentes amostras. A proteína cristal produzida pela subespécie B. thuringiensis var. israelensis foi purificada por ultracentrifugação e utilizada para imunizar coelhas e produzir soros hiperimunes. Tais soros foram posteriormente usados para avaliar o nível de proteína cristal em bioinseticidas comerciais e em culturas de laboratório desta bactéria utilizando-se a técnica do imunodot. Os resultados foram obtidos por comparação de reações com concentrações conhecidas de proteína cristal permitindo assim avaliar com segurança os níveis desta proteína em várias preparações.

The development of the production and use of Bacillus thuringiensis in Brazil at a commercial scale faces certain difficulties, among them the establishment of efficient methodologies for the quantitation of toxic products to be commercialized. Presently, the amount of toxin is given in percentage by analyzing the samples total protein content. Such methodology however, does not measure the actual amount of active protein present in the product, since most strains express different endotoxin genes and might even produce b-toxin. Since the various types of toxins exhibit different antigenic characteristics, this work has as objective the utilization of fast immunological techniques to quantify the level of crystal protein. Crystal protein produced by a subspecies of Bacillus thuringiensis var. israelensis was purified by ultracentrifugation and utilized to immunize rabbits and to produce hiperimmune sera. Such sera were latter used to evaluate the level of proteins on commercial bioinsecticide and on laboratory cultures of B. thuringiensis through the immunodot technique. The results were obtained by comparison of data obtained from reactions with known concentrations of crystal protein permitting to evaluate the level of such protein on various materials.