Genetic variability and physiological traits of Saccharomyces cerevisiae strains isolated from "Vale dos Vinhedos" vineyards reflect agricultural practices and history of this Brazilian wet subtropical area.

Vale dos Vinhedos appellation of origin has a very recent history as industrial wine making region. In this study we investigated the genetic and phenotypic variability of Saccharomyces cerevisiae strains isolated from South-Brazilian vineyards in order to evaluate strain fermentation aptitude and copper and sulphites tolerance. Merlot grape bunches were collected from three vineyards and yeast isolation was performed after single bunch fermentation. High genotypic variability was found and most of the genotypes revealed to be vine-specific. No industrial strain dissemination was present in the sampled vineyards, although it has been wildly reported in traditional winemaking countries. From the phenotypic traits analysis these Brazilian native strains showed good fermentation performances, good tolerance to sulphites and, in particular, a high copper tolerance level. Copper is the most important metal in the formulation of fungicides against downy mildew (Plasmopara viticola), one of the most harmful disease of the vines, and other fungal pests. The high tolerance to copper suggests an environmental adaptation to the strong use of copper-based fungicides, requested by the wet subtropical climate.

Indica rice cultivar IRGA 424, transformed with cry genes of B. thuringiensis, provided high resistance against Spodoptera frugiperda (Lepidoptera: Noctuidae).

Plant expression of the entomopathogenic bacteria Bacillus thuringiensis cry gene has reduced the damage created by insect pests in several economically important cultures. For this study, we have conducted genetic transformation of the indica rice "IRGA 424", via Agrobacterium tumefaciens, using the B. thuringiensis cry1Aa and cry1B genes, with the objective of obtaining rice plants resistant to the insect pests from this culture. The gene constructions harbor the promoters maize proteinase inhibitor and ubiquitin. The results showed that high concentration of the hormone 2,4-dichlorophenoxyacetic acid and agarose as the gelling agent helped the production of embryogenic calli for the analyzed cultivar. More than 80% of the obtained transformed plants revealed the integration, using polymerase chain reaction, of the cry1Aa and cry1B genes. Analysis of the expression of the heterologous protein by Western blotting revealed the expression of the Cry1B delta-endotoxin in IRGA 424 plants transformed with the ubiquitin promoter. Data showed the production and dissemination of a high number of embryogenic calli in addition to obtaining plants transformed with the cry1Aa and cry1B genes until the reproductive phase. The feed bioassays with the transformed plants and Spodoptera frugiperda (JE Smith) larvae indicated high rates of mortality to the insect target. The highest corrected mortality rate achieved under laboratory conditions with Bt-rice plants transformed with the cry1B and cry1Aa genes was 94 and 84%, respectively. Thus, our results demonstrated the great potential of transformed Bt-rice plants in controlling the damage caused by these insect pests in rice paddy fields.

PCR and bioassays screening of Bacillus thuringiensis isolates from rice-fields of Rio Grande do Sul, specific to lepidopterans and coleopterans

Visando a selecão 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 Agar Nutriente durante 12 h e submetidos a extracão de DNA total. Os fragmentos amplificados foram analisados em géis de agarose (1-1,5 per center). Os resultados referentes ao total de isolados selecionados mostraram que 56,51 per center foram potencialmente específicos a lepidópteros (cry1, cry2 e cry9) e 21,73 per center a coleópteros (cry3 e cry7/8), tendo sua distribuicão 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 per center), o qual havia sido pré-selecionado pela presenca 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 per center de mortalidade às larvas de Oryzophagus oryzae, tendo esses sido pré-selecionados pela presenca de genes cry3. Os resultados dos bioensaios confirmam a predicão da atividade de Bt por PCR, a qual deve estar diretamente relacionada aos genes cry que codificam as proteínas inseticidas específicas.

Distribuição de genes cry de Bacillus thuringiensis isolados de solos do Estado do Rio Grande do Sul, Brasil / Distribution of cry genes of Bacillus thuringiensis isolated from soils of the State of Rio Grande do Sul, Brazil

A bactéria Bacillus thuringiensis (Bt) caracteriza-se pela produçäo de proteínas tóxicas a representantes de diversas ordens de insetos, as quais säo 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 säo limitados no Rio Grande do Sul, essa pesquisa objetivou avaliar a distribuiçäo 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 por cento. A presença de genes cry9 foi detectada em 47,82 por cento dos isolados, seguido de cry3 (15,21 por cento), cry1 e cry7 (ambos com 6,52 por cento) e cry2 (2,17 por cento). Oito perfis genéticos foram identificados, sendo o perfil cry9 (39,13 por cento) 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 poderäo ser avaliadas quanto à toxicidade in vivo contra insetos-praga das plantas cultivadas