ABSTRACT The endophytic bacteria belong to a larger group of microorganisms that have their life-cycle partly or entirely inside the plant and are located in intra and inter-cellular spaces or in the vascular tissue. These bacteria can be found colonizing aerial parts or roots. This review aims to analyze the colonization strategies of endophytic bacteria through interaction with plants, as well as to highlight the metabolic influence of these organisms in plant tissues, which result in physiological and biochemical changes. Depending on the different mechanisms used internally to colonize a plant, these microorganisms are called obligate, facultative, or passive endophytes. Phytostimulation, biofertilization and biological control are mechanisms that result in the development of the plant through the production of plant hormones, bioavailability of nutrients and antagonistic action to phytopathogens, respectively. The association between endophytic bacteria and plants features important benefits such as significant increases in growth, plant biomass, length of roots, dry matter production, and grain yield. Studies show that there is a great diversity of endophytic bacteria colonizing plant structures that result in several benefits to the host plant.

ABSTRACT The microbial composition of different types,in ecosystems (including agro-ecosystems), has been investigated in a rapidly growing number of studies in the past few years. The importance of microorganisms, regarding the maintenance and stability of nutrients in agroecosystems, is a key to maintain the sustainability of a crop. Molecular tools to study microbial communities are possible through many methods such as RISA, DGGE, TGGE, clone libraries, T-RFLP, RAPD, SSCP and more recently NGS (Next-Generation Sequencing). DGGE is widely employed to characterize the diversity and the community dynamics of microorganisms in the environment, making possible to find out specific groups through functional genes, allowing access to data that cannot be obtained by cultural methods. The aim of this paper is to review the functional groups related to agroecosystems and to indicate the critical choice of DNA primers pairs and targeted DNA regions that may be used in PCR-based methods such as the DGGE technique in order to evaluate the microbial communities in a variety of environments.

This work presents bibliographic data on the role and function of microbial diversity. The increasing use of probiotics and prebiotics foods has led to the studies on their actual functions in the human body. It is known that in the environment, microorganisms are extremely important in recycling of nutrients, balance of trophic chains, vital physiological activities in the plants and animals, as well as the conservation of natural habitats. In human food, these microscopic organisms contribute from flavoring products to the synthesis of antimicrobial substances and vitamins essential to living beings.

RESUMO Esta publicação apresenta alguns dados sobre as interações entre as bactérias e as plantas hospedeiras, considerando os simbiontes e os patógenos das plantas de importância agrícola, com ênfase nos agroecossistemas de arroz irrigado. Neste trabalho foram abordados aspectos relacionados a essas bactérias endofíticas, tais como: (i) endofíticos e suas interações com as plantas cultivadas; (ii) rizobactérias promotoras do crescimento vegetal [plant growth-promoting rhizobacteria (PGPR)] e sua aplicabilidade no controle biológico; (iii) bactérias endofíticas versus PGPR no controle biológico de fitopatógenos; (iv) aplicação de recombinantes endofíticos na agrobiotecnologia. No final do artigo estão apresentadas as opiniões e perspectivas dos autores sobre as bactérias que vivem associadas às plantas cultivadas nos agroecossistemas.

ABSTRACT This paper presents some data on the interactions of the bacteria and the host plants, considering the symbionts and pathogens of plants of agricultural importance, focusing on rice fields. This article addresses issues related to these endophytic bacteria, such as: (i) endophytes and their interactions with the cultivated plants; (ii) promoting rhizobacteria plant growth [plant growth-promoting rhizobacteria (PGPR)] and its application in biological control; (iii) endophytic bacteria PGPR against the biological control of plant pathogens; (iv) application of endophytic recombinant in agricultural biotechnology. This paper ends show the point of view and perspectives of the authors about bacteria that live associated with the plants grown in agroecosystems.

Oryzophagus oryzae (Costa Lima 1936) (Coleoptera: Curculionidae) and Spodoptera frugiperda (J E Smith, 1797) (Lepidoptera: Noctuidae) cause important crop losses in southern Brazil. Control is possible by the use of the bacteria Bacillus thuringiensis and extracts of Melia azedarach. This study aimed to evaluate the mortality, in vivo, of O. oryzae and S. frugiperda submitted to two isolates of B. thuringiensis and the aqueous extract of M. azedarach. The LC50 for O. oryzae due to bacteria was 5.40μg/mL (Bt 2014-2) and due to plant extract 0.90μg/mL. For S. frugiperda, the Bt 1958-2 bacterial suspension (1.10(10)UFC/mL) caused a 100% of corrected mortality, showing that the purified Cry proteins caused a CL10 of 268μg/mL five days after the treatments, and M. azedarach toxins caused a CL50 173μg/mL four days after the treatment. Corrected mortality for O. oryzae and S. frugiperda in the interaction between the bacterial and plant toxins were 11 and 6%, respectively. In the PCR analysis of B. thuringiensis isolates, DNA fragments were enlarged and corresponded to the cry1 and cry2 genes for Bt 1958-2. Thus, it could be concluded that the usage of Bt 2014-2 active against O. oryzae larvae; Bt 1958-2 for S. frugiperda and, for both the insect species, M. azedarach aqueous extract could be used.

Efeito tóxico de óleos essenciais de plantas medicinais em adultos de Sitophilus oryzae (Linnaeus) (Coleoptera, Curculionidae). Grãos armazenados estão sujeitos a perdas na qualidade física, sanitária e nutricional, desde sua maturação fisiológica até o momento do consumo. Uma alternativa para o manejo de pragas são os inseticidas botânicos. O presente trabalho teve como objetivo avaliar o efeito inseticida de óleos essenciais de Cymbopogon citratus (folha), Zingiber officinale (raiz) e Mentha sp. (folha), obtidos através do método de hidrodestilação,no controle de adultos do gorgulho-do-arroz, S. oryzae. A caracterização química dos compostos encontrados nos óleos essenciais foi realizada por cromatografia gasosa, sendo observados como principais componentes para C. citratus: geranial (48%) e neral (31%), para Z. officinale: 16% de geranial, 13% de α-zingibereno, 10% de neral e 5% de α-farneseno e para Mentha sp.: 92% de mentol. Os bioensaios foram realizados por aplicação tópica e fumigação. Os resultados da aplicação tópica mostraram que o óleo essencial de C. citratus apresenta maior toxicidade quando comparado à Mentha sp.e ao Z. officinale, pois apresentou uma CL50 de 0.027 µL mL-2 e um menor período de exposição, causando 70% e 100% de mortalidade de adultos de S. oryzae, respectivamente, 24 e 48 horas após dos tratamentos. Nos ensaios com fumigação, em adultos de S. oryzae, o óleo essencial de Z. officinale, apresentou a menor CL50 (1,18 µL cm-2) e o menor período de exposição, causando 70% de mortalidade em 24 horas após a aplicação dos tratamentos. Desse modo conclui-se que os óleos de C. citratus e Z. officinale podem ser recomendados no controle de adultos do gorgulho-do-arroz S. oryzae.

Toxic effects of essential plant oils in adult Sitophilus oryzae (Linnaeus) (Coleoptera, Curculionidae). Stored grains are subject to losses in quality nutritional value and in sanitation from the time they are stored to the time they are consumed. Botanical insecticides may offer an alternative solution for pest control. The objective was to test the insecticidal properties of the essential oils of Cymbopogon citratus (leaf), Zingiber officinale (root) and Mentha sp. (leaf). The efficacy of these oils was tested to control the rice weevil, S. oryzae, using hydrodistillation. Chemical analysis of the essential oils was carried out by gas chromatography. Major components of C. citratus were geranial (48%) and neral (31%), of Z. officinale were α-zingibereno (13%), geranial (16%), neral (10%) and α-farneseno (5%) and of Mentha sp. was menthol (92%). Bioassays were carried out by fumigation and topical application. In topical application assays, the essential oil of C. citratus had greater toxicity (LC50 0.027 µL mL-1) and shorter exposure time than the oils of the other two plants. After 24 h and 48 h, 70% and 100% mortality of S. oryzae occurred, respectively. In fumigation assays, essential oil of Z. officinale had a lower LC50 (1.18 µL cm-2) and 70% mortality after 24 h exposure. Therefore, we recommend the use of essential oils of C. citratus and Z. officinale to control the rice weevil S. oryzae.

Bacilos entomopatogênicos são bactérias observadas principalmente no solo, consideradas importantes agentes no controle biológico de doenças e pragas da agricultura. No entanto, existem poucos estudos que relatam a influência das práticas agrícolas nas populações de Bacillus spp. Essa pesquisa objetivou avaliar a freqüência de bactérias entomopatogênicas pertencentes ao gênero Bacillus obtidas de amostras de solo coletadas no ano agrícola de 2006/2007 de diferentes sistemas de cultivo de arroz irrigado da EEA-IRGA, Cachoeirinha (RS). Foram analisadas 36 amostras de solos, oriundas de três sistemas de plantio de arroz irrigado, pré-germinado (SPG), convencional (SPC), plantio direto (SPD) e de uma área sem plantio de arroz (C). Para tanto, um grama de solo foi diluído, pasteurizado e inoculado em Ágar Nutriente (AN), incubado a 30 °C, por 24 horas. Todas as unidades formadoras de colônias bacterianas (UFC) foram repicadas em meios seletivos e as espécies, B. thuringiensis, B. sphaericus e B. cereus, foram identificadas por microscopia de contraste de fase. Os resultados revelaram que não houve diferença significativa entre os sistemas de plantio analisados. Por outro lado, a frequência de Bacillus spp. variou entre as diferentes fases do plantio de arroz nos períodos amostrados. A irrigação foi o principal fator que contribuiu para a variação da frequência dos Bacillus em sistemas de plantio de arroz irrigado da EEA-IRGA.

Entomopathogenic Bacillus are bacteria found mainly in the soil and are considered important agents in biological control of agricultural diseases and pests. However, there are fen studies describing the influence that different agricultural practices have on Bacillus spp. populations. This study proposes to determine the occurrence frequency of entomopathogenic bacteria from Bacillus genus in soil samples. These were collected during the agricultural year 2006/07 from different irrigated rice cultivation systems used in the EEA-IRGA plantations in Cachoerinha, State of Rio Grande do Sul, Brazil. We analyzed 36 soil samples taken from three irrigated rice plantation systems; (i) Pre-germinated (SPG); (ii) Conventional (SPC); and (iii) Direct plantation (SPD) and a fourth sample from a non-planted area (C). In each test one gram of soil was diluted, pasteurized and inoculated in Nutrient Agar (NA) and incubated for 24 hours at 30°C. All the bacteria colony-forming units (CFU) were transplanted to selective mediums and the B. thuringiensis, B. sphaericus and B. cereus species were identified using a phase contrast microscope. No significant differences in the occurrence frequencies were found in the soil samples of all plantation systems analyzed. On the other hand, the Bacillus spp frequency varied in the sampled period during the different growing phases of the irrigated rice in the EEA-IRGA plantations evaluated. Irrigation was the main factor contributing to variation of the Bacillus in the plantations.

As cepas e proteínas Cry1Ab e Cry1Ac sintetizadas por Bacillus thuringiensis thuringiensis e B. thuringiensis kurstaki, foram avaliadas nos fitopatógenos: Rhizoctonia solani,Pyricularia grisea,Fusarium oxysporum e F. solani, os quais tiveram seu crescimento micelial reduzido após a incubação na presença das cepas bacterianas. Em relação às proteínas Cry, não houve formação de halo de inibição nas concentrações avaliadas.

Cry1Ab and Cry1Ac strains and proteins synthesized by Bacillus thuringiensis thuringiensis and B. thuringiensis kurstaki were assessed in the following phytopathogens: Rhizoctonia solani,Pyricularia grisea,Fusarium oxysporum and F. solani, which had their micelial growth decreased after incubation in the presence of the bacterial strains. As to Cry proteins, there were no inhibition halo development in the assessed concentrations.

Spodoptera frugiperda (J. E. Smith, 1797) (Lepidoptera, Noctuidae) está entre os insetos-praga que mais danificam culturas, sendo possível seu controle com produtos à base de Bacillus thuringiensis Berliner, 1911 (Bt). Um dos principais parasitóides de S. frugiperda é Campoletis flavicincta (Ashmead, 1890) (Hymenoptera, Ichneumonidae). A interação de Bt aizawai e de C. flavicincta através da análise histopatológica do ovo do parasitóide em lagartas de S. frugiperda infectadas foi avaliada. Lotes de lagartas de segundo ínstar foram analisados: 1) parasitadas e infectadas por Bt aizawai (Xentari/Abbott Laboratórios do Brasil Ltda) a 0,5mg ml-1 e 2) apenas parasitadas por C. flavicincta (controle). Todas as lagartas foram fixadas em "Bouin Hollande", em intervalos de 6, 12 e 24 horas posteriores à infecção. Após o preparo das lagartas em blocos de parafina, foram efetuados cortes histológicos de 6µm de espessura. A análise histológica de lagartas parasitadas e infectadas indicou não ter havido alteração no ovo do parasitóide, resultante da ação do bacilo. Esse resultado indica que ovos de C. flavicincta não são afetados por Bt aizawai no interior do corpo de lagartas de S. frugiperda.

Spodoptera frugiperda (J. E. Smith, 1797) (Lepidoptera, Noctuidae) is one of the most important insect pests that affect several crops, and it can be controled with Bacillus thuringiensis Berliner, 1911 (Bt) products. One of the main parasitoids of S. frugiperda is Campoletis flavicincta (Ashmead, 1890) (Hymenoptera, Ichneumonidae). The interaction between Bt aizawai and C. flavicincta by histopathological analysis of the egg of parasitoid in S. frugiperda infected larvae was evaluated. Groups of second instar larvae were analyzed: 1) parasitized and infected by Bt aizawai (Xentari /Abbott Laboratórios do Brasil Ltda) at 0.5mg ml-1 and 2) only parasitized by C. flavicincta (control). All the larvae were fixed in "Bouin Hollande", at intervals of 6, 12 and 24 hours after the infection. After the preparation of the larvae in paraffin blocks, histological cuts of 6µm thickness were made. The histological analysis of parasitized and infected larvae indicated no alteration in the egg of the parasitoid, as a result of the action of the bacillus. This result suggests that eggs of C. flavicincta are not affected by Bt aizawai inside of the body of larvae of S. frugiperda.

A Anticarsia gemmatalis é responsável pelo uso de inseticidas químicos na cultura da soja, ocasionando um significativo aumento nos custos das lavouras e um grande desequilíbrio no ecossistema. O uso de agentes microbianos, como Bacillus thuringiensis sorovar kurstaki (Btk) e Vírus de Poliedrose Nuclear de Anticarsia gemmatalis (VPNAg), é uma alternativa para o controle químico de insetos-praga. Na análise da interação de bactérias e vírus entomopatogênicos, considera-se importante o modo de ação in vitro desses agentes de controle microbiano. Assim, o presente trabalho objetiva a análise histopatológica do sistema digestivo das lagartas de A. gemmatalis, após a interação dos entomopatógenos Btk e VPNAg, representados nas formulações Dipel e Baculovirus anticarsia, respectivamente. As avaliações foram realizadas com lagartas de 2º ínstar, onde a mortalidade foi avaliada diariamente, e a histopatologia foi realizada com lagartas coletadas nos períodos de 1, 3, 6, 12 e 24 horas após a aplicação dos tratamentos. Os resultados dos ensaios in vivo, revelam que o tratamento referente à associação VPNAg/Btk (98.68% de mortalidade) foi mais eficiente que VPNAg (81.28% de mortalidade) isoladamente, porém o Btk isoladamente causou 100% de mortalidade. Os tratamentos mostraram diferenças significativas (P <0,05) entre AgNPV e Btk,AgNPV e AgNPV/Btk. As análises de patogenicidade do VPNAg e Btk em lagartas de A. gemmatalis sugerem que os produtos Dipel e Baculovirus anticarsia foram mais eficientes, quando utilizados simultaneamente, pois a ação do VPNAg foi intensificada quando utilizada em associação com Btk, provocando alterações no intestino médio das lagartas a partir de 6 horas após os tratamentos. Quando os entomopatógenos foram utilizados isoladamente, as alterações das células intestinais foram observadas apenas 12 horas após a aplicação dos tratamentos.

The Anticarsia gemmatalis is responsible for the use of chemical insecticides in the soybean culture, causing a significant increase in the costs of farming and a great unbalance in the ecosystem. The use of microbial agents, like Bacillus thuringiensis serovar kurstaki (Btk) and Anticarsia gemmatalis nucleopolyhedrovirus (AgNPV), they are an alternative to chemical control of the pest insects. In the interaction analysis of the entomopathogenic bacteria and virus it is considered important the in vitro action mode of these microbiology control agents. Therefore, the present study aims the histopathological analysis of the A. gemmatalis larvae digestive system after the interaction in vivo of the entomopathogenic Btk and AgNPV, represented the Dipel and Baculovirus anticarsia formulations, respectively. The evaluations were realized in larvae of 2nd instar, in which the mortality was evaluated daily, and a histopathology was done with collected larvae in time of 1, 3, 6, 12 and 24 hours after the treatments application. The results of the in vivo assays reveal that the treatment using the association of AgNPV-Btk (98.68% of mortality) was more efficient than using AgNPV isolatedly (81.28% of mortality), but the Btk when used isolatedly had a mortality of 100%. The treatments showed significant (P<0.05) differences between AgNPV and Btk,AgNPV and AgNPV/Btk. The histopathological analysis of the AgNPV and Btk in A. gemmatalis larvae suggests that the Dipel and Baculovirus anticarsia products were more efficient when they were used simultaneously, because the action of AgNPV was intensified when used in association with Btk, causing changes in the larvae midgut after 6 hours of treatments. When the entomopathogens were used isolating the gut cells alterations were observed only 12 hours after the treatments.

O Rio Grande do Sul é um dos maiores produtores de milho do país, porém pouco se conhece em termos de inimigos naturais dos principais insetos-praga da cultura. O objetivo do presente trabalho foi realizar um levantamento dos parasitóides da lagarta-do-cartucho do milho, Spodoptera frugiperda. Para tanto, foram efetuadas amostragens em áreas de lavouras de milho do Instituto Riograndense do Arroz (IRGA), em Cachoeirinha, RS, nos anos agrícolas 1999/2000 e 2000/2001. A coleta dos insetos foi realizada semanalmente, durante o ciclo da cultura. As lagartas foram individualizadas e mantidas em laboratório até a emergência dos adultos ou dos parasitóides. Foram coletadas 1425 lagartas no primeiro período de amostragens e 518 no segundo. Destas, 18,11% e 22,01% encontravam-se parasitadas, com a presença dos seguintes parasitóides: Chelonus sp., Cotesia sp. e Exaticolus sp. (Hym., Braconidae), Campoletis flavicincta e Ophion sp. (Hym., Ichneumonidae) e Archytas incertus e Lespesia archippivora (Dip., Tachinidae). Houve predomínio de C. flavicincta, correspondendo a 63,57% e 76,32% dos parasitóides, respectivamente para o primeiro e segundo período de amostragens. Em C. flavicincta, ocorreu hiperparasitismo por Conura sp. (Chalcididae) e Aphanogmus sp. (Ceraphronidae).

Rio Grande do Sul State is one of the greatest maize producers of the country, however, little is known in terms of the main natural enemies associated to insect pests in this crop. The objective of this work was to survey parasitoids of fall-armyworm, Spodoptera frugiperda. Samplings in maize areas of the Instituto Riograndense do Arroz (IRGA) were conducted in Cachoeirinha, RS, during the growth seasons of 1999/2000 and 2000/2001. Insect larvae were collected weekly, during the crop cycle. Larvae were individualized and kept in laboratory until emergence of adults or parasitoids. In the first and second crop seasons, 1425 and 518 larvae were collected, respectively. Out of these, 18.11% and 22.01% were parasitized, with the following parasitoids being observed: Chelonus sp., Cotesia sp. and Exaticolus sp. (Hym., Braconidae), Campoletis flavicincta and Ophion sp. (Hym., Ichneumonidae), and Archytas incertus and Lespesia archippivora (Dip., Tachinidae). There was predominance of C. flavicincta, corresponding to 63.57% and 76.32% of the parasitoids, respectively, for the first and second seasons. In C. flavicincta hiperparasitism occurred by Conura sp. (Chalcididae) and Aphanogmus sp. (Ceraphronidae).

Visando a seleção de seis grupos de genes cry de Bacillus thuringiensis (Bt), que codificam proteínas ativas para coleópteros e lepidópteros pragas do arroz, 46 isolados de Bt provenientes de amostras de solos das regiões orizícolas do Rio Grande do Sul (RS), foram testados por PCR. Os isolados de Bt foram crescidos em Ágar Nutriente durante 12 h e submetidos a extração de DNA total. Os fragmentos amplificados foram analisados em géis de agarose (1-1,5%). Os resultados referentes ao total de isolados selecionados mostraram que 56,51% foram potencialmente específicos a lepidópteros (cry1, cry2 e cry9) e 21,73% a coleópteros (cry3 e cry7/8), tendo sua distribuição homogênea entre as regiões orizícolas do RS. Apenas os genes cry2 foram localizados somente na região Litoral. Nos bioensaios com lagartas de Spodoptera frugiperda o isolado Bt 2027-1 obteve a maior mortalidade corrigida (25%), o qual havia sido pré-selecionado pela presença de genes cry9. Para a mesma espécie, os testes de toxicidade através de proteínas purificadas de Bt aizawai HD68 revelaram uma DL50 de 0,95 mg/larva. Dois isolados de Bt causaram 100% de mortalidade às larvas de Oryzophagus oryzae, tendo esses sido pré-selecionados pela presença de genes cry3. Os resultados dos bioensaios confirmam a predição da atividade de Bt por PCR, a qual deve estar diretamente relacionada aos genes cry que codificam as proteínas inseticidas específicas.

Bacillus thuringiensis (Bt) isolates from soil samples of rice-fields in Rio Grande do Sul (RS) were tested through PCR, aiming at the screening of six groups of Bt cry genes, which codify active proteins for coleopterans and lepidopterans rice pests, and their bioinsecticide potential regarding their use in IPM system, as well. Forty six Bt isolates were grown in Agar Nutrient for 12 h and submitted to total DNA extraction. The amplified fragments were analyzed in agarose gel (1-1.5%). The screening isolates showed that 56.51% were potentially lepidopterans specific (cry1, cry2 and cry9) and 21.73% were coleopteran specific (cry3 and cry7/8), with a homogeneous distribution in the rice-field areas in Rio Grande do Sul. Cry2 genes were found just once and in the Litoral area. Bioassays against Spodoptera frugiperda larvae showed the highest corrected mortality (25%) with Bt 2027-1 isolate selected by the presence of cry9 genes. The toxicity bioassays carried out with S. frugiperda using purified proteins of Bt aizawai HD68 indicated a LD50 of 0.95 µg/larvae. Two Bt isolates carrying the cry3 genes (PCR detection) caused a 100% mortality to Oryzophagus oryzae larvae. Bioassay results confirmed the prediction of Bt activity by PCR, which must have a straight relationship with the cry genes that codify those specific insecticidal proteins.

Os insetos constituem uma das principais causas de danos à produção agrícola no mundo. O controle de insetos tem sido realizado por meio de agroquímicos e, em muito menor escala, pelo emprego de inseticidas biológicos. As plantas transgênicas resistentes a insetos representam uma nova alternativa no combate aos insetos-praga das lavouras. A bactéria entomopatogênica Bacillus thuringiensis Berlinier (Bt) é a fonte dos genes de resistência nas chamadas plantas-Bt, produzidas comercialmente. No presente trabalho de revisão, são abordados os aspectos relacionados à bactéria Bt como fonte de genes de resistência a insetos-pragas, plantas geneticamente modificadas, vantagens do uso de plantas-Bt, bem como perspectivas dessa ferramenta biotecnológica.

Insect pests are a major worldwide cause of damage to agriculture. The control of insect pests was primarily relied on agrochemical and, in a much smaller scale, on biological insecticides. Insect-resistant transgenic plants represent a new alternative to the protection of crops against insect pests. The entomopathogenic bacterium Bacillus thuringiensis (Bt) is the source of resistance genes for the commercial production of the so called Bt-plants. In this review, the main features of Bt as source of resistance genes will be described as well as the main aspects of insect-resistant transgenic plants and advantages of using Bt-plants.

A bactéria Bacillus thuringiensis (Bt) caracteriza-se pela produção de proteínas tóxicas a representantes de diversas ordens de insetos, as quais são codificadas por genes cry. Devido a esta característica, mais de 40.000 cepas de Bt foram isoladas e cerca de 190 genes cry, caracterizados. Como os dados sobre Bt são limitados no Rio Grande do Sul, essa pesquisa objetivou avaliar a distribuição de seis famílias de genes cry de Bt, desse estado, que codificam proteínas ativas contra insetos-praga. O perfil dos 46 isolados de solos do Rio Grande do Sul foi avaliado, por PCR com os primers que detectam os genes cry1, cry2, cry3, cry7 cry8 e cry9 e suas respectivas proteínas foram analisadas por SDS-PAGE a 10%. A presença de genes cry9 foi detectada em 47,82% dos isolados, seguido de cry3 (15,21%), cry1 e cry7 (ambos com 6,52%) e cry2 (2,17%). Oito perfis genéticos foram identificados, sendo o perfil cry9 (39,13%) o mais freqüente. A análise protéica de Bt identificou 14 famílias de proteínas Cry possivelmente codificadas por genes presentes nos isolados, além de proteínas desconhecidas que podem caracterizar novos genes cry. Esses isolados revelam a presença de genes que codificam proteínas específicas contra lepidópteros e coleópteros, as quais poderão ser avaliadas quanto à toxicidade in vivo contra insetos-praga das plantas cultivadas.

The Bacillus thuringiensis (Bt) bacterium is characterized by the production of toxic protein to representatives of several insect orders, which are coded by cry genes. Due to this characteristic, more than 40.000 Bt strains were isolated and around 190 cry genes characterized. As the data on Bt are limited in Rio Grande do Sul, this research aimed the evaluation of the distribution of six Bt families of cry genes, in this state, that codify active proteins against insect-pest. The 46 isolates profiles of soil samples from Rio Grande do Sul were evaluated, by PCR with primers that detect cry1, cry2, cry3, cry7, cry8 and cry9 genes, and its respective proteins were analyzed by SDS-PAGE 10%. The presence of cry9 genes were detected in 47.82% of the isolates, followed by cry3 (15.21%), cry1 and cry7 (both with 6.52%) and cry2 (2.17%). Eight genetic profiles were identified, being the profile cry9 (39.13%) the most frequent one. The Bt proteic analysis identified 14 families of Cry protein that are possibly coded by the genes in these isolates, besides the unknown proteins that can characterize new cry genes. These isolates reveal the presence of genes that codify specific proteins against lepidopterans and coleopterans, which can be evaluated for its in vivo toxicity against insect-pest of the cultivated plants.

A bactéria Bacillus thuringiensis (Bt) é caracterizada pela sua habilidade de produção de corpos paraesporais durante a esporulação. As proteínas cry1 têm atividade inseticida e são altamente específicas para certos insetos e não são tóxicas para outros insetos, plantas e vertebrados. Neste trabalho, a patogenicidade de doze isolados foram testados contra Anticarsia gemmatalis, um dos mais importantes insetos pragas da cultura da soja. Para tanto, foi aplicada sobre a superfície da dieta uma suspensão de esporo-cristal e a mortalidade de lagartas de Anticarsia gemmatalis foi avaliada sete dias após a aplicação. Quando comparado com uma linhagem de Bt controle, conhecido pela sua alta toxicidade para lagartas da soja, quatro novos isolados exibiram atividade tóxica superior, acima de 90% de mortalidade. Foi utilizada a técnica de PCR para amplificar fragmentos de DNA de regiões codificantes de genes cry1. Os isolados de Bt com alta mortalidade produziram produtos de PCR de tamanho esperado, em torno de 280 pb, isolados não tóxicos ou pouco tóxico não produziram qualquer produto de PCR ou mostraram fragmentos amplificados com padrões diferentes do esperado. Os isolados de Bt com alta atividade contra a lagarta da soja mostraram a presença de proteínas com tamanho de aproximadamente 150 kDa, quando o extrato protéico total foi analisado em SDS-PAGE.

The bacterium Bacillus thuringiensis (Bt) is characterized by its ability to produce proteic crystalline inclusions during sporulation. Cry1 protein has insecticidal activity and is highly specific to certain insects and not toxic to unrelated insects, plants or vertebrates. In this work, the patogenicity of twelve Bt isolates was tested against Anticarsia gemmatalis, one of the most important insect pests of soybeans. Spore-crystal complex was applied to the surface of artificial diets and the mortality of A. gemmatalis larvae was assessed seven days after each treatment. When compared to a control Bt isolate known by its high toxicity to A. gemmatalis larvae, four novel Bt isolates exhibited even higher toxic activities against the insect, resulting in more than 90% mortality. PCR was used to amplify DNA fragments related to known cry1 genes. Bt strains with high toxicity produced expected PCR products of around 280 bp, whereas non-toxic or low toxic strains did not produce any PCR product or showed amplified fragments of different sizes. Toxic Bt isolates also exhibited an expected protein profile when total protein extracts were evaluated by SDS-PAGE.