Compatibility of Beauveria bassiana and alternative phytosanitary products.

AIMS: To assess the compatibility of alternative phytosanitary products (APPs) with the entomopathogenic fungus Beauveria bassiana (Bals.) Vuillemin. METHODS AND RESULTS: The APPs used were Baicao, Orobor® , Topneem, Rotenat and Compostonat at concentrations recommended by the manufacturers (CR), and then half (1/2CR) and double (2CR). To evaluate the compatibility, the following parameters were analysed: germination, colony forming units (CFUs), vegetative growth and conidia production. Compatibility was assessed using rates of toxicity and biological index. The association of APPs and B. bassiana was also evaluated for pathogenicity against larvae of third instar Anagasta kuehniella Zeller (Lepidoptera: Pyralidae) (1·0 × 108 conidia per ml). The germination of conidia of B. bassiana was reduced only by the product Orobor® . The CFUs were not affected by any products. The colony diameter was reduced when in contact with the products Baicao and Orobor® . For conidia production per colony, all products reduced this parameter. None of the products affected the pathogenicity of the fungus on A. kuehniella. CONCLUSIONS: All APPs tested are compatible with B. bassiana. SIGNIFICANCE AND IMPACT OF THE STUDY: The natural products tested and used in the field are considered compatible with B. bassiana under laboratory conditions, making possible new studies and the use of these in insect control.

Action of natural phytosanitary products on Bacillus thuringiensis subsp. kurstaki S-1905.

The objective of this study was to evaluate the effects of natural phytosanitary products (NPs) on spores and crystals of Bacillus thuringiensis subsp. kurstaki S-1905 (Btk S-1905). For the spore assay, NPs and bacteria were applied in combination and individually. For the combined application, Btk S-1905 + NP mixtures were inoculated on nutrient agar (NA), and for the separate applications, the NPs were spread on NA plates, which were later inoculated with the pathogen. The number of colony-forming units (CFU) per milliliter was quantified after 18 h of incubation. For the crystal protein degradation assay, the Btk S-1905 + NP mixtures were added to the diet of Anticarsia gemmatalis (Lepidoptera: Erebidae), and mortality was evaluated at the following time points: 12, 24, 48, and 72 h. Scanning electron microscopy and agarose gel electrophoresis were carried out. Biogermex and Ecolife® reduced the CFU ml-1 in both combined and separate applications. Biogermex, Ecolife®, and Planta Clean were antagonistic to the action of bacterial toxins, and no product affected the morphology or resulted in the degradation of the crystal proteins. The remaining products evaluated did not reduce the CFU ml-1 and had additive effect when combined with the crystal toxin.

Potential Side Effects of the Entomopathogenic Fungus Metarhizium anisopliae on the Egg Parasitoid Trichogramma pretiosum (Hymenoptera: Trichogrammatidae) Under Controlled Conditions.

The purpose was to evaluate the side effects of strains Metarhizium anisopliae (Metsch.) Sorokin sensu lato Unioeste 43 and M. anisopliae sensu stricto ESALQ 1641 on Trichogramma pretiosum Riley (Hymenoptera: Trichogrammatidae) under controlled conditions. A free-choice test for parasitism was performed, with the confinement of T. pretiosum females mated with cards (1 × 5 cm) containing age-standardized Anagasta kuehniella Zeller eggs, either sprayed with a fungal strain (109 conidia/ml) or 0.01% v/v Tween 80 (control). For the no-choice tests, T. pretiosum females mated were confined with cards sprayed with fungal strains before or after parasitism, and cards with fungal applications at different times. The number of parasitized eggs, percentage of emergence, longevity, egg-to-adult period, sex ratio, total and confirmed mortality by the fungus, and longevity of females that parasitized previously sprayed eggs, were assessed. Histological analysis of immature phases was also performed. The fungus was repellent to T. pretiosum in the free-choice test, while in the no-choice test, fungal applications before and after parasitism did not affect the number of eggs parasitized by T. pretiosum or the sex ratio of emerging adults. However, both strains affected adult emergence rates, the egg-to-adult period, and longevity. Overall, both M. anisopliae strains had minor effects on these biological parameters of T. pretiosum under controlled conditions. Hyphae were not detected in histological observations of immature stages of the parasitoid.

Técnicas para avaliação in vitro do efeito de herbicidas sobre bacillus thuringiensis berliner var. kurstaki

ABSTRACT The objective of this research was to compare techniques for the in-vitro evaluation of the effect of herbicides onBacillus thuringiensis Berliner var. kurstaki (Btk). The herbicides sulfentrazone, lactofen, imazethapyr, fenoxaprop and metribuzin were tested at two concentrations, on cells and spores, in nutrient agar (NA) solid medium (SM), and in nutrient broth (NB) liquid medium (LM), in addition to their metabolization (MT) by Btk. For cells, the fungicides in NA were tested incorporated into the medium and on filter-paper disks impregnated with the products. Evaluation was made, respectively, of the number of colony-unit formers per milliliter CFU.mL-1 and of the formation of an inhibition halo. For cells, the Btk + products + NB mixture was incubated in Erlenmeyer flasks (30 ± 2° C, 150rpm, 24h) and then the NB was diluted and inoculated onto NA, followed by evaluation of the number of CFU.mL-1. The MT and spore tests were performed in liquid medium (water for MT and NB for spores). The effects of the products varied with their concentration, the evaluation technique used and the developmental stage of the microorganism. The LM evaluation technique proved to be the best for evaluating the effect of herbicides on Btk cells, as it did not show any diffusion limitations and it also allowed for quantification. The herbicides, regardless of their concentration, were incompatible with the cells, although sulfetrazone and imazethapyr were metabolized by Btk. All herbicides were compatible with the spores, allowing for germination, but the concentration and time influenced the results.

RESUMO Objetivou-se comparar técnicas para a avaliação in vitro do efeito de herbicidas sobre Bacillus thuringiensis Berliner var. kurstaki (Btk). Foram testados os herbicidas sulfentrazone, lactofen, imazethapyr, fenoxaprop e metribuzin em duas concentrações, sobre células e esporos, em meio ágar nutriente sólido (MS) e caldo nutriente (CN) - líquido (ML), além da metabolização (MT) de tais produtos por Btk. Para células em MS, os herbicidas foram testados incorporados ao meio e em discos de papel-filtro impregnados nos produtos. Avaliou-se, respectivamente, as unidades formadoras de colônia por mililitro (UFC/mL) e a formação de halo de inibição. Para células, a mistura Btk + produtos + CN foi incubada em frascos erlemmeyer (30 ± 2° C, 150 rpm, 24h) e, em seguida, a mistura foi diluída e inoculada em MS, avaliando-se o número de UFC/mL. Os testes de MT e esporos foram realizados em líquido (água para MT e CN para esporos). O efeito dos produtos variou conforme a concentração utilizada, técnica de avaliação e a fase/estágio do microrganismo, sendo a técnica de avaliação em ML a mais indicada para a avaliação do efeito dos herbicidas sobre células de Btk, pois não apresenta limitações para difusão e permite quantificação. Os herbicidas, independentemente da concentração, foram incompatíveis para a célula, porém sulfetrazone e imazethapyr foram metabolizados por Btk. Todos herbicidas foram compatíveis com esporos, possibilitando a germinação, porém a concentração e tempo influenciaram nos resultados.

Entomopatógenos associados a lagartas de bombyx mori l. (lepidoptera: bombycidae) no estado do paraná

ABSTRACT The state of Paraná exceeds all other Brazilian states in terms of silk production. Approximately 7,000 agricultural producers engage in this activity, in about 200 counties. During the caterpillar stage, silkworms (Bombyx mori L.) are susceptible to a number of diseases caused by viruses, fungi, and bacteria. These diseases seriously impact cocoon production, and consequently result in losses for producers. To study the entomopathogens associated to B. mori caterpillars in the state of Paraná, the present study was carried out at the main producing regions from November 2002 to April 2003. Live and/or dead caterpillars suspected of infection were collected directly from the rearing huts of silk-producing companies. The collected material was stored in plastic containers, properly identified and placed in cooler boxes. In the laboratory, the caterpillars were processed by specific methodologies according to the suspected pathogen. After isolation, the entomopathogens Baculovirus (31.4%), Beauveria bassiana (Bals.) Vuill. (5.3%) and Bacillus thuringiensis Berliner (2.2%) were identified.

RESUMO O Estado do Paraná é o maior produtor sericícola brasileiro. A atividade é explorada em aproximadamente 200 municípios, envolvendo cerca de 7.000 produtores rurais. Durante o estágio de lagarta, o bicho-da-seda (Bombyx mori L.) é susceptível a doenças causadas por vírus, fungos e bactérias. Estas doenças podem gerar sérios danos na produção de casulos e, conseqüentemente, prejuízos para o produtor. Com o objetivo de se verificar os entomopatógenos associados a lagartas de B. mori no Estado do Paraná, foi desenvolvido o presente trabalho nas principais regiões produtoras, no período de novembro de 2002 a abril de 2003. Para tanto, lagartas suspeitas de infecção, vivas e/ou mortas foram coletadas diretamente dos barracões de criação das empresas sericícolas. O material coletado foi armazenado em frascos plásticos identificados e colocado em caixas térmico. No laboratório, as lagartas foram processadas utilizando-se metodologias específicas de acordo com o patógeno suspeito. Após o isolamento foram identificados os entomopatógenos Baculovírus (31,4%), Beauveriabassiana (Bals.) Vuill. (5,3%) e Bacillus thuringiensis Berliner (2,2%).

Técnicas para avaliação do efeito in vitro de fungicidas sobre bacillus thuringiensis var. kurstaki

ABSTRACT The aim of this study was to compare techniques for the invitro evaluation of the effect of fungicides on Bacillus thuringiensis var. kurstaki (Btk). The fungicides epoxiconazole, fentin hydroxide, azoxystrobin, tebuconazole, and carbendazim were tested at two concentrations, on cells and spores, in nutrient agar (NA) solid medium (SM), and in nutrient broth (NB) liquid medium (LM), with an evaluation also being made of their metabolization (MT) by Btk. For cells, the fungicides in NA were tested incorporated into the medium and on filterpaper disks. The evaluation consisted, respectively, of the number of CFU/mL and of the formation of an inhibition halo. For cells, the Bt+products + NB mixture was incubated in Erlenmeyer flasks (30 ± 2° C, 150rpm, 24h) and then was diluted and inoculated onto NA; the number of CFU/mL was evaluated. The MT and spore tests were performed in liquid medium (water). The effects of the products varied with their concentration, technique used, contact time, and developmental stage of the pathogen. The LM test is the most recommendable for evaluating the effect on cells. It was observed that most fungicides were compatible with cells, and only epoxiconazole and azoxystrobin were metabolized. Fentin hydroxide was the only fungicide incompatible with spores.

RESUMO Objetivou-se comparar técnicas para a avaliação in vitro do efeito de fungicidas sobre Bacillus thuringiensis var. kurstaki (Btk). Foram testados os fungicidas epoxiconazole, fentin hidróxido, azoxystrobin, tebuconazole e carbendazin em duas concentrações, sobre células e esporos, em ágar nutriente (AN)-sólido (MS) e caldo nutriente (CN)-líquido (ML), além da metabolização (MT) destes por Btk. Para células em AN, os fungicidas foram testados incorporados ao meio e em discos de papel-filtro. Avaliou-se, respectivamente, as UFC/mL e a formação de halo de inibição. Para células, a mistura Bt + produtos + CN foi incubada em erlemmeyers (30 ± 2° C, 150 rpm, 24h), e em seguida, diluida e inoculada em AN, avaliandose o número de UFC/mL. Os testes de MT e esporos foram realizados em líquido (água). O efeito dos produtos variou conforme a concentração, técnica, tempo de contato e fase/estágio do desenvolvimento do patógeno. O teste ML é o mais indicado para a avaliação do efeito em células, sendo observado que a maioria dos fungicidas mostrou-se compatível com as mesmas, e apenas epoxiconazole e azoxystrobin foram metabolizados. Fentin hidróxido foi o único incompatível com esporos.