Desenvolvimento de micoinseticidas: uma abordagem multidisciplinar visando o controle biológico do percevejo bronzeado do eucalipto Thaumastocoris peregrinus (Hemiptera: Thaumastocoridae)
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Data
2018-12-07
Autores
Velozo, Simone Graziele Moio
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Universidade Estadual Paulista (Unesp)
Resumo
Thaumastocoris peregrinus Carpintero & Dellapé, 2006 (Hemiptera: Thaumastocoridae), originário da Austrália e com ampla distribuição geográfica é responsável por perdas econômicas na cultura do eucalipto em várias regiões no mundo. Devido ao hábito alimentar deste inseto em várias espécies de Eucalyptus, este é classificado como uma praga de importância mundial. A busca por controle biológico de T. peregrinus, aliada à investigação de métodos para produção de micopesticidas bem como a necessidade de maior entendimento de fungos entomopatogênicos para controle de uma praga florestal, nos objetivou em estudar: seleção de isolados e avaliação da patogenicidade ao T. peregrinus; caracterização de Metarhizium anisopliae e Beauveria bassiana em diferentes temperaturas; fermentação sólida e líquida destes fungos nos biorreatores JKI-Solid Fermenter System, Laboratory Fermenter Prophyta L03® (M. anisopliae) e Minifors Infors HT® (B. bassiana) com diferentes escalas de produção; colheita de conídios puros oriundos de fermentação sólida; otimização dos processos de produção através de secagem em sistema estéril durante a produção do fungo; colheita e estabilidade de M. anisopliae através da secagem automatizada e caracterização de conídios puros dispersos em líquidos (Tween® 0,1% e óleo) quanto ao processo de sedimentação acelerada e distribuição do tamanho de partículas. Metarhizium anisopliae, B. bassiana, Cordyceps spp. e Sporothrix insectorum são fungos entomopatogênicos com alto potencial para controlar T. peregrinus, com destaque para M. anisopliae, o mais virulento. As temperaturas de 25 e 30 °C são ideais para a ação dos fungos B. bassiana (IBCB227), M. anisopliae (IBCB425) e Cordyceps farinosa (IBCB220) controlando T. peregrinus. Temperaturas acima de 37 °C podem causar a inativação dos isolados IBCB425 e IBCB227, sendo a condição ideal de 20 a 30 °C e de 15 a 30 °C, respectivamente. Na fase de produção, o biorreator JKI-Solid Fermenter System foi eficaz na produção de IBCB425 e IBCB227 em substratos sólidos com um rendimento máximo de aproximadamente 6,0 x 109 e 1,0 x 109 conídios/g de massa seca de fungo+substrato após 14 dias de fermentação, respectivamente, entretanto, a colheita de conídios secos só foi possível para IBCB425, devido ao não desprendimento dos esporos de IBCB227 no equipamento de colheita. Por esta razão, o Laboratory Fermenter Prophyta L03® foi utilizado para produzir grandes quantidade de conídios do isolado IBCB425 e obteve um rendimento de 6,4 x 109 conídios/g de massa seca de fungo+substrato. O Mycoharvester® versão 5b colheu e separou, eficazmente, partículas de M. anisopliae, entretanto, recomenda-se a secagem do substrato por 7 dias com uma taxa de aeração de 2L/min visando uma maior estabilidade do conídio a 25 °C. Na fase de formulação, o adjuvante Nimbus® apresentou efeito negativo na germinação dos conídios independentemente da combinação em que este adjuvante estava, resultando de 0 a 13% de germinação. As combinações Óleo de girassol + Natur’óleo® (40%), Óleo de milho + Iharol Gold® (30%) e Óleo de milho + Assist® (30%) apresentaram germinação de 86,55; 89,66 e 97,55%, respectivamente, sendo as taxas mais altas de germinação dentre as formulações. As formulações testadas foram mais eficientes no controle de adultos de T. peregrinus, sendo que a formulação Óleo de girassol + Natur’óleo® (40%) apresentou uma maior mortalidade. Além disso, os conídios de M. anisopliae apresentava melhores resultados quando em contato com o óleo, tanto para a distribuição do tamanho de partículas quanto para o processo de sedimentação acelerada. A fermentação líquida é outra possibilidade de produzir IBCB227 e IBCB425, onde meios de cultura a base de arroz têm um rendimento de 4,6 x 108 e 5,5 x 107 esporos/mL, respectivamente. Beauveria bassiana pode ter um rendimento aproximado de até 1 x 109 esporos/mL em biorreatores automatizados, sendo o consumo de glicose diretamente relacionado à capacidade de esporulação e produção de biomassa. Este estudo possibilitou a compreensão de várias etapas para produção de produtos biológicos a base de fungos, como multiplicação, colheita e aplicação desses produtos para controle de T. peregrinus, além disso, os métodos podem ser transferidos para outros insetos praga.
Thaumastocoris peregrinus Carpintero & Dellapé, 2006 (Hemiptera: Thaumastocoridae), originated in Australia and has a wide geographic distribution is responsible for economic losses in the eucalyptus plantations in several regions in the world. Due to the feeding behavior of this insect on several species of Eucalyptus, this is classified as a pest of world importance. The search for biological control of T. peregrinus, together with the investigation of methods to produce myopesticides as well as the need for a better understanding of entomopathogenic fungi to control a forest pest, aimed to study: selection of isolates and evaluation of pathogenicity to T. peregrinus; characterization of Metarhizium anisopliae and Beauveria bassiana at different temperatures; solid and liquid fermentation of these fungi in the JKI-Solid Fermenter System, Laboratory Fermenter Prophyta L03® (M. anisopliae) and Minifors Infors HT® (B. bassiana) bioreactors with different production scales; harvesting of pure conidia from solid fermentation; optimization of the production processes through drying in a sterile system during fungus production; harvesting and stability of M. anisopliae by automated drying and characterization of the pure conidia of this fungus dispersed in liquids (Tween® 0.1% and oil) in the process of accelerated sedimentation and distribution of particle size. Metarhizium anisopliae, B. bassiana, Cordyceps spp. and Sporothrix insectorum are entomopathogenic fungi with high potential to control T. peregrinus, with emphasis on M. anisopliae, the most virulent. Temperatures of 25 and 30 °C are the best for the fungi B. bassiana (IBCB227), M. anisopliae (IBCB425) and Cordyceps farinosa (IBCB220) controlling T. peregrinus. Temperatures above 37 °C can cause inactivation of IBCB425 and IBCB227 isolates, the ideal condition being 20-30 °C and 15-30 °C, respectively. In the production phase, the JKI-Solid Fermenter System bioreactor was effective in the production of IBCB425 and IBCB227 on solid substrates with a maximum yield of approximately 6.0 x 109 and 1.0 x 109 conidia/g of fungus+substrate dry mass after 14 days of respectively, however, the collection of dry conidia was only possible for IBCB425, due to the non-shedding of IBCB227 spores in the harvesting equipment. For this reason, Laboratory Fermenter Prophyta L03® was used to produce large conidia of IBCB425 isolate and yielded 6.4 x 109 conidia/g of fungus+substrate dry mass. Mycoharvester® version 5b efficiently harvested and separated M. anisopliae particles, however, drying of the substrate for 7 days at an aeration rate of 2L/min is recommended for greater conidial stability at 25 °C. In the formulation phase, the Nimbus® adjuvant had a negative effect on conidia germination regardless of the combination in which this adjuvant was present, resulting in 0 to 13% germination. The combination of sunflower oil + Natur'oleo® (40%), Corn oil + Iharol Gold® (30%) and Maize oil + Assist® (30%) presented germination of 86.55; 89.66 and 97.55%, respectively, being the highest germination rates among the formulations. The formulations tested were more efficient in the control of adults of T. peregrinus, and the formulation Sunflower oil + Natur'óleo ® (40%) had a higher mortality. In addition, the conidia of M. anisopliae presented better results when in contact with the oil, both for the particle size distribution and for the accelerated sedimentation process. Liquid fermentation is another possibility of producing IBCB227 and IBCB425, where rice-based culture media has a yield of 4.6 x 108 and 5.5 x 107 spores/mL, respectively. Beauveria bassiana can achieve an approximate yield of up to 1.0 x 109 spores/mL in automated bioreactors, and glucose consumption is directly related to sporulation capacity and biomass production. This study allowed the understanding of several stages for the production of biological products based on fungi, such as multiplication, harvest and application of these products to control T. peregrinus, in addition, the methods can be transferred to other insect pests.
Thaumastocoris peregrinus Carpintero & Dellapé, 2006 (Hemiptera: Thaumastocoridae), originated in Australia and has a wide geographic distribution is responsible for economic losses in the eucalyptus plantations in several regions in the world. Due to the feeding behavior of this insect on several species of Eucalyptus, this is classified as a pest of world importance. The search for biological control of T. peregrinus, together with the investigation of methods to produce myopesticides as well as the need for a better understanding of entomopathogenic fungi to control a forest pest, aimed to study: selection of isolates and evaluation of pathogenicity to T. peregrinus; characterization of Metarhizium anisopliae and Beauveria bassiana at different temperatures; solid and liquid fermentation of these fungi in the JKI-Solid Fermenter System, Laboratory Fermenter Prophyta L03® (M. anisopliae) and Minifors Infors HT® (B. bassiana) bioreactors with different production scales; harvesting of pure conidia from solid fermentation; optimization of the production processes through drying in a sterile system during fungus production; harvesting and stability of M. anisopliae by automated drying and characterization of the pure conidia of this fungus dispersed in liquids (Tween® 0.1% and oil) in the process of accelerated sedimentation and distribution of particle size. Metarhizium anisopliae, B. bassiana, Cordyceps spp. and Sporothrix insectorum are entomopathogenic fungi with high potential to control T. peregrinus, with emphasis on M. anisopliae, the most virulent. Temperatures of 25 and 30 °C are the best for the fungi B. bassiana (IBCB227), M. anisopliae (IBCB425) and Cordyceps farinosa (IBCB220) controlling T. peregrinus. Temperatures above 37 °C can cause inactivation of IBCB425 and IBCB227 isolates, the ideal condition being 20-30 °C and 15-30 °C, respectively. In the production phase, the JKI-Solid Fermenter System bioreactor was effective in the production of IBCB425 and IBCB227 on solid substrates with a maximum yield of approximately 6.0 x 109 and 1.0 x 109 conidia/g of fungus+substrate dry mass after 14 days of respectively, however, the collection of dry conidia was only possible for IBCB425, due to the non-shedding of IBCB227 spores in the harvesting equipment. For this reason, Laboratory Fermenter Prophyta L03® was used to produce large conidia of IBCB425 isolate and yielded 6.4 x 109 conidia/g of fungus+substrate dry mass. Mycoharvester® version 5b efficiently harvested and separated M. anisopliae particles, however, drying of the substrate for 7 days at an aeration rate of 2L/min is recommended for greater conidial stability at 25 °C. In the formulation phase, the Nimbus® adjuvant had a negative effect on conidia germination regardless of the combination in which this adjuvant was present, resulting in 0 to 13% germination. The combination of sunflower oil + Natur'oleo® (40%), Corn oil + Iharol Gold® (30%) and Maize oil + Assist® (30%) presented germination of 86.55; 89.66 and 97.55%, respectively, being the highest germination rates among the formulations. The formulations tested were more efficient in the control of adults of T. peregrinus, and the formulation Sunflower oil + Natur'óleo ® (40%) had a higher mortality. In addition, the conidia of M. anisopliae presented better results when in contact with the oil, both for the particle size distribution and for the accelerated sedimentation process. Liquid fermentation is another possibility of producing IBCB227 and IBCB425, where rice-based culture media has a yield of 4.6 x 108 and 5.5 x 107 spores/mL, respectively. Beauveria bassiana can achieve an approximate yield of up to 1.0 x 109 spores/mL in automated bioreactors, and glucose consumption is directly related to sporulation capacity and biomass production. This study allowed the understanding of several stages for the production of biological products based on fungi, such as multiplication, harvest and application of these products to control T. peregrinus, in addition, the methods can be transferred to other insect pests.
Descrição
Palavras-chave
percevejo bronzeado, fungos entomopatogênicos, fermentação em estado sólido, fermentação submersa, produção de biopesticidas, bronze bug, entomopathogenic fungi, solid state fermentation, submerged fermentation, biopesticide production