Effect of entomopathogenic fungi on the control of the coffee berry borer in the laboratory

Among the management and control tactics of the coffee berry borer Hypothenemus hampei (Ferrari), there is the use of entomopathogenic fungi. Due to the importance of prospecting isolates of entomopathogenic fungi for the control of the coffee berry borer, the objective of this study was to evaluate the efficiency of 26 isolates of entomopathogenic fungi in the control of this insect pest in the laboratory. The coffee berry borers were immersed in a solution adjusted to the concentration of 1 to 3 × 108 conidia/mL of each isolate and the control treatment (sterilized water). After seven days total mortality and confirmed mortality were evaluated. The isolates that caused the highest mortality and two commercial isolates were selected for evaluation of lethal concentration (LC50 and LC90) and lethal time (LT50 and LT90). Coffee berry borers were treated at different conidia concentrations for lethal concentration to assess total and confirmed mortality. For a lethal time, the coffee berry borers were treated at the concentration of 108 conidia/mL of the selected isolates, and, after two days and every 24 hours until the eighth day, the number of dead individuals was verified. Among the 26 isolates evaluated, 24 presented mortality higher than the control treatment, and three presented mortality higher than 85%. In the LC50 and LC90 assays, the IBCB 353 and IBCB 364 isolates were more lethal to H. hampei. In the LT50 and LT90 assays, the IBCB 66 and IBCB 353 isolates caused lethality in a shorter time.

Caenorhabditis elegans as an indicator of toxicity of Bacillus thuringiensis strains to Meloidogyne incognita race 3 / Caenorhabditis elegans como indicador de toxicidade de estirpes de Bacillus thuringiensis a Meloidogyne incognita raça 3

ABSTRACT: The cotton plant (Gossypium hirsutum) is affected by several diseases of economic importance, among them root-knot nematode (Meloidogyne incognita races 3 and 4). Methods to control this disease include the application of nematicides, solarization, deep plowing, crop rotation and use of antagonistic microorganisms. Among species of Bacillus, there are strains that act as bioregulators and antagonists of several pathogens. Tests to identify these strains are hampered by the difficulty of obtaining large populations of the pathogen and by the time of execution of the in vivo tests that should be conducted for about 90 days. The objective of this research was to compare the toxicity of B. thuringiensis strains to two nematodes, M. incognita and Caenorhabditis elegans, evaluating the possibility of using C. elegans as an indicator for the selection of strains with biocontrol potential against M. incognita. Therefore, the toxicity of nine B. thuringiensis strains on C. elegans and M. incognita was evaluated under laboratory and greenhouse conditions. Most strains toxic to C. elegans in vitro were also toxic to M. incognita, and three of them (S906, S1192, S2036) significantly reduced the populations of the two nematodes. The toxic effect of B. thuringiensis strains on C. elegans was like that reported for the same bacterial isolates on M. incognita in vivo. These results suggested that it is plausible to use C. elegans as an indicator of toxicity for selection of B. thuringiensis strains toxic to M. incognita.

RESUMO: O algodoeiro (Gossypium hirsutum) é acometido por várias doenças de importância econômica, dentre as quais a meloidoginose (Meloidogyne incognita raças 3 e 4). Entre os métodos de controle dessa doença, destacam-se as aplicações de nematicidas, a solarização, a aração profunda, a rotação de culturas e o uso de microrganismos antagonistas. Dentre as espécies do gênero Bacillus, existem estirpes que atuam como biorreguladores e antagonistas de vários patógenos. Os testes para identificação dessas estirpes são prejudicados pela dificuldade de se obter grandes populações do patógeno e pelo tempo de execução dos testes in vivo que devem ser conduzidos por cerca de 90 dias. Diante disso, o presente trabalho teve como objetivo comparar a toxicidade de estirpes de B. thuringiensis a dois nematoides, M. icognita e Caenorhabditis elegans, verificando a possibilidade de empregar C. elegans como indicador para a seleção de estirpes com potencial de biocontrole contra M. incognita. Para tanto, a toxicidade de nove estirpes de B. thuringiensis para C. elegans e M. incognita foi avaliada em laboratório e em casa de vegetação. A maioria das estirpes tóxicas ao C. elegans in vitro, também foi tóxica ao M. incognita, sendo que três delas (S906, S1192, S2036) reduziram significativamente as populações dos dois nematoides. O efeito tóxico apresentado pelas estirpes de B. thuringiensis contra C. elegans foram similares aos apresentados pelos mesmos isolados contra M. incognita in vivo. Esses resultados sugerem que é plausível o uso do C. elegans como indicador de toxicidade para seleção de estirpes de B. thuringiensis tóxicas a M. incognita.

Evidence of field-evolved resistance of Spodoptera frugiperda to Bt corn expressing Cry1F in Brazil that is still sensitive to modified Bt toxins.

Brazil ranked second only to the United States in hectares planted to genetically modified crops in 2013. Recently corn producers in the Cerrado region reported that the control of Spodoptera frugiperda with Bt corn expressing Cry1Fa has decreased, forcing them to use chemicals to reduce the damage caused by this insect pest. A colony of S. frugiperda was established from individuals collected in 2013 from Cry1Fa corn plants (SfBt) in Brazil and shown to have at least more than ten-fold higher resistance levels compared with a susceptible colony (Sflab). Laboratory assays on corn leaves showed that in contrast to SfLab population, the SfBt larvae were able to survive by feeding on Cry1Fa corn leaves. The SfBt population was maintained without selection for eight generations and shown to maintain high levels of resistance to Cry1Fa toxin. SfBt showed higher cross-resistance to Cry1Aa than to Cry1Ab or Cry1Ac toxins. As previously reported, Cry1A toxins competed the binding of Cry1Fa to brush border membrane vesicles (BBMV) from SfLab insects, explaining cross-resistance to Cry1A toxins. In contrast Cry2A toxins did not compete Cry1Fa binding to SfLab-BBMV and no cross-resistance to Cry2A was observed, although Cry2A toxins show low toxicity to S. frugiperda. Bioassays with Cry1AbMod and Cry1AcMod show that they are highly active against both the SfLab and the SfBt populations. The bioassay data reported here show that insects collected from Cry1Fa corn in the Cerrado region were resistant to Cry1Fa suggesting that resistance contributed to field failures of Cry1Fa corn to control S. frugiperda.

Synergistic activity of Bacillus thuringiensis toxins against Simulium spp. larvae.

Species of Simulium spread diseases in humans and animals such as onchocerciasis and mansonelosis, causing health problems and economic loses. One alternative for controlling these insects is the use of Bacillus thuringiensis serovar israelensis (Bti). This bacterium produces different dipteran-active Cry and Cyt toxins and has been widely used in blackfly biological control programs worldwide. Studies on other insect targets have revealed the role of individual Cry and Cyt proteins in toxicity and demonstrated a synergistic effect among them. However, the insecticidal activity and interactions of these proteins against Simulium larvae have not been reported. In this study we demonstrate that Cry4Ba is the most effective toxin followed by Cry4Aa and Cry11Aa. Cry10Aa and Cyt1Aa were not toxic when administered alone but both were able to synergise the activity of Cry4B and Cry11Aa toxins. Cyt1Aa is also able to synergise with Cry4Aa. The mixture of all toxin-producing strains showed the greatest level of synergism, but still lower than the Bti parental strain.

Translocation and insecticidal activity of Bacillus thuringiensis living inside of plants.

The major biological pesticide for the control of insect infestations of crops, Bacillus thuringiensis was found to be present naturally within cotton plants from fields that had never been treated with commercial formulations of this bacterium. The ability of B. thuringiensis to colonize plants as an endophyte was further established by the introduction of a strain marked by production of green fluorescent protein (GFP). After inoculation of this preparation close to the roots of cotton and cabbage seedlings, GFP-marked bacteria could be re-isolated from all parts of the plant, having entered the roots and migrated through the xylem. Leaves taken from the treated plants were able to cause toxicity when fed to the Lepidoptera Spodoptera frugiperda (cotton) and Plutella xylostella (cabbage). These results open up new horizons for understanding the natural ecology and evolution of B. thuringiensis and use of B. thuringiensis in insect control.